Engineering Drawing Short Questions

Define engineering drawing. Why drawing is called universal language of engineers? Ans1:-A drawing drawn by an engineer having engineering knowledge for the drawing purposes is an engineering drawing. It is meant for communicating his ideas, thoughts and designs to others. Engineering drawing is a starting point of all engineering branches such as Mechanical, Production, Civil, Electrical, Electronics, Computer science, Chemical etc. It is spoken, read, and written in its own way.Engineering drawing has its own grammar in the theory of projections, its idioms in conventional practices, its punctuations in the types of lines, its abbreviations, symbols and its descriptions in the constructions. Q2 – Name different types of drawing instruments. Ans2 – Drawing board, T-square, Set Square, Scales, Pencil and sand paper block, Drawing pins or cello-tape, Duster or handkerchief, eraser etc. Q3 – Why pencil is rotated in finger while drawing a long line? Ans3 – T he pencil is rotated in finger while drawing a long line in order to get a line of uniform thickness throughout.Q4 – How will you test the set square and T-square? Ans4 – Testing of T-square – (i) Check all screw heads and tighten, if necessary (ii) In order to check the T-square, first of all draw a horizontal line. Now reverse the T-square and again draw a horizontal line with working edge. If both the lines coincide with each other, then the working edge of Tsquare is alright. If there is a difference in two lines, then working edge is not correct and the line gives twice the error of the working edge. This error should be rectified by scraping the edge with a scraper or a sharp knife.Testing of set-squares – The straightness of edges of the set-square can be checked by drawing a vertical line. Then reverse the set-square and draw again vertical line. If there is any difference between the two vertical lines then working edge is not correct and the lin e gives twice the error. This error can be removed by straightening the edges by means of a scraper or sand paper. Q5 – What are the standard sizes of drawing sheets according to I. S. I. and which is suitable for drawing work? Ans5 – The standard size of sheets according to I. S. I. are A0(1189 X 841), A1 841 X 594), A2(594 X 420), A3(420 X 297), A4(297 X 210) and A5(210 X 148). Drawing sheet of size 594 X 420 i. e. A2 size is generally used by engineering students as it is very handy and easy for drawing work in class. Q6 – What are the ways of sharpening a pencil for good and accurate work and which type of pencil is more suitable for drawing work? Ans6 – There are two ways of sharpening a pencil (i) a small piece of sand paper of zero grade, pasted upon a piece of wood. (ii) Sharpeners. Usually hard pencils such as H, 2H etc are used for making the engineering drawing.Q7 – Why cello-tape is used instead of drawing pins, now a day? Ans7 – Now a days, cello tapes are used in place of drawing pins for its practical convenience as the drafter, Tsquare and set-squares can be moved easily over the tape. Q8 – What is layout of drawing sheet? Ans8 – The selection of suitable scale and allotment of proper space for margin, title block, parts list, revision panel, folding marks etc. on the drawing sheet is known as layout of drawing sheet. Q9 – Why is the layout of sheet is necessary? Ans9 – Layout of the drawing on the drawing sheet is necessary in order to make its reading easy and speedy.The title blocks, parts list etc will provide all the required information. Q10 – List out the contents of title block and material list Ans10 – The title block should contain at least the following informations. (i) Name of the institution (ii) Name of title of drawing (iii) Name, Class and Roll no. of the student (iv) Scale (v) Drawing number (vi) Symbols denoting the method of projection Q11 â €“ What is the necessity of folding a drawing print? Ans11 – Folding marks are made on the sheet to facilitate folding of prints for the purposes of filing and binding in the proper and easy manner.Q12 – What do you mean by convention or code? Ans12 – The representation of any matter by some sign or mark on the drawing is known as convention or code. The conventions make the drawing simple and easy to draw. Q13 – What do you understand by thickness of lines? Ans13 – There are three distinct thickness of lines used in engineering drawing. These lines are specified as thick, medium and thin lines. The line specified as thick is usually 3 times thicker and the line specified as medium is 2 times thicker than a thin line. Q14 – Where and why a cutting plane is drawn in a drawing?Ans14 – The section plane are generally perpendicular planes. The projection of a section plane, to which it is perpendicular, is a straight line. This line w ill be parallel, perpendicular or inclined to the x-y line. The cutting plane is drawn in a drawing to show the inner details of an object. Q15 – What is the necessity of convention breaks and convention of materials? Ans15 – Long members of uniform cross-section such as rods, shafts, pipes etc. are generally shown in the middle by the conventional breaks so as to accommodate their view of whole length on the drawing sheet without reducing the scale.The exact length of the member is shown by the dimension. Q16 – Why the conventional representation of common features are adopted on the drawing? Ans16 – The conventional representation of common features are adopted on the drawing to save the unnecessary time or space on the drawing. Q17 – What are the main requirements of lettering? Ans17:- 1) The knowledge of shape and proportion of each letter. 2) The knowledge of the order and direction of the strokes used in making letters. 3) The knowledge of th e general composition of letters. 4) The knowledge of rules for combining letters into words and words into sentences.Q18 – What is lettering? Ans18 – The art of writing the alphabets A, B, C,†¦Ã¢â‚¬ ¦. Z and numbers such as 1, 2, 3†¦Ã¢â‚¬ ¦0 etc. is known as lettering. Q19 – What do you mean by composition of letters? Ans19 – The composition means the composing of letters into words and words into sentences. The letters are so arranged that the open area between two letters of a word appears equal to the eye judgement. Q20 – What do you mean by uniformity of letters? Ans20 – The uniformity of lettering means keeping the height, inclination, spacing and strength of letters to be same. It is very essential for good lettering in engineering drawing.Q21 – What do you mean by normal, compressed and extended lettering? Ans21 – Normal lettering: – The normal lettering have normal height and width and are used for gen eral purposes. The width of the normal letter is about 0. 67 times of the height of the letter. Compressed lettering: – The compressed lettering are those which are written in the narrow space. These are used when the space is limited. The widths of the condensed letters are less than height. Extended lettering: – The extended lettering are those which are wider than noramal letters but of the same height.Q22 – What are the guidelines and why they are necessary in lettering? Ans22:- The lines which are used to regulate the height and inclination to the letters and numerals are known as guidelines. These are to be drawn at random. The guidelines are used to regulate the uniformity of the letters. Q23 – What do you mean by single stroke letters? Ans 23:- Single stroke letters means that the thickness of the line of the letter should be such as is obtained in one stroke of the pencil. Single stroke letters are of two types. 1) Vertical 2) Inclined (75deg. Wi th horizontal) Q24 – What is the gothic and roman lettering?Ans24 – Gothic lettering – The lettering in which all the alphabets are of uniform width or thickness is known as gothic lettering. It can be divided into following groups. (i) Vertical or Upright vertical gothic lettering (ii) Inclined or Italic gothic lettering Roman lettering – The lettering in which all the alphabets are composed of thick and thin elements is known as roman lettering and can either be vertical or inclined. Q25 – What do you mean by freehand lettering? Ans25 – The art of writing the alphabets without the use of drawing instrument is called freehand lettering. The freehand lettering is of the following types. a) Vertical or upright freehand gothic lettering. (i) Single stroke vertical freehand gothic lettering. (ii) Lowercase vertical freehand gothic lettering. (b) Inclined or italic freehand gothic lettering. (iii) Single stroke italic freehand gothic lettering. (iv) Lower case italic freehand gothic lettering. Q26 – What should be the grade of pencil used for lettering? Ans26 – HB and H grade pencils sharpened to a conical point should be used for lettering. To keep the stroke of the letters uniform, the pencils should be rotated between the thumb and fingers while lettering. Hard pencils such as 2H or 3H should be used to draw guidelines.Q27 – What is the importance of dimensioning? Ans27:-1) Dimensioning expresses all the sizes and other information necessary to define the object. 2) It must be done with due regard to manufacturing processes and inspection requirements. 3) The dimensioning also includes expression of tolerances necessary for the correct functioning of the part given to be assembled. Q28 – What is dimensioning? Ans28 – The art of writing the various sizes or measurements on the finished drawing of an object is known as dimensioning. Q29 – What do you understand by the term notatio n of dimensioning? Ans29 –The notation of dimensioning consists of dimension lines, extension lines, arrow heads, dimension figures, notes, symbols etc. Q30 – What is a leader or pointer line? How a leader should be drawn? Ans30 – A leader is a thin continuous line drawn from note of the figure to show where it applies. It is terminated by an arrow head or a dot. The arrow head touches the outline, whereas the dot is placed within the outline of the object. The leader is generally drawn at any convenient angle, usually 30? , 45? , and 60? but not less than 30?. Q31 – Explain with the help of a simple sketch (i) size dimensions (ii) location dimensions.Ans31 – Size dimension – The dimensions which indicate the various sizes of the object such as length, breadth, diameter etc. are known as size dimensions. These dimensions are represented by letter ‘S’. Figure. Location dimension – The dimensions which locate the position o f one feature w. r. t. the other feature are known as location dimensions. Distances between the centre lines of the holes from the edges are given by location dimensions. These dimensions are marked by letter ‘L’. Figure. Q32 – What are the aligned system and unidirectional system of dimensioning? OrWhat are the different methods of dimensioning? Ans32:-1) Aligned Method: – In aligned system, the dimensions shall be placed parallel to and above the dimension lines, preferably in the middle and not by interrupting the dimension lines. Here the dimensions can be read from the bottom or from the right side of the drawing. Figure. 2) Unidirectional Method: – In this system dimensions shall be horizontally placed so that they can be read from the bottom of the drawing sheet. Here the dimension lines may be interrupted preferably near the middle for the insertion of dimensions. Figure.Q33 – What are the general rules of dimensioning? Ans33:-1) Eve ry dimension must be given, but no single dimension should be repeated. 2) Dimensions should be placed outside the views. 3) Avoid dimensioning to hidden lines wherever possible. 4) Dimension lines should not cross any other line of the drawing. 5) Aligned system of dimensioning is recommended. Q34 – Explain with simple sketches, the methods of dimensioning (i) circles (ii) radii (iii) angles (iv) spherical shapes (v) holes. Ans34 – Q35 – Explain with the help of sketches (i) chain dimensioning (ii) parallel dimensioning and (iii) combined dimensioning.Ans35 – Chain Dimensioning – In this system, dimensions are arranged in a straight line. Figure. Parallel dimensioning – In this arrangement, all the dimensions are given from common base line. The smaller dimensions are placed nearer the view and the larger further away so that the extension lines do not cross dimensions lines. Figure. Combined dimensioning – Combined dimensioning is t he result of the simultaneous use of chain and parallel dimensioning. Figure. Q36 – What is a scale? Ans36:-A scale is defined as the proportion by which we either reduce or increases the actual size of the object on a drawing. ) Full size scale:-The scale in which the actual measurements of the object are drawn to same size on the drawing is known as full size scale. 2) Reducing scale: – The scale in which the actual measurements of the object are reduced to some proportion is known as reducing scale. 3) Enlarging scale: – The scale in which the actual measurements of the object are increased to some proportion is known as enlarging scale. Q37 – What is the representative fraction (R. F. ) or scale factor (S. F. )? Ans37:-The ratio of the drawing size of an object to its actual size is called the Representative fraction.R. F. = Dimension of an object on sheet / Actual dimension of an object Q38 – What are the main uses of scale? Ans38 – The following are the main uses of scale in engineering practice. (i) The scales are used to prepare reduced or enlarged size drawings. (ii) The scales are used to set off dimensions. (iii) The scales are used to measure distances directly. Q39 – What are the information necessary for scale? Ans39 – To construct a scale, the following informations are necessary. (i) The representative fraction (R. F. ) of the scale. (ii) The units to be presented either in metric or British measures. iii) The maximum length of the scale. Q40 – What is difference between plane scale and diagonal scale? Ans40:-Plain Scale:-A plain scale is simply a line which is divided into a suitable number of equal parts, the first of which is further sub-divided into small parts. It is used to represent either two units or a unit and its fraction such as km and hm, m and dm, etc. Diagonal Scale:- A diagonal scale is used when very minute distances such as 0. 1 mm etc. are to be accurately measure d or when measurements are required in three units; for example dm, cm, and mm. Q41 – What is the principle of a diagonal scale?Ans41: – The principle of diagonal scale is to divide a short line into any number of equal parts by following the diagonal division’s method of construction. Q42 – What is the difference between a quadrilateral and a polygon? Ans42 – Quadrilateral – A quadrilateral is a plane figure bounded by four straight lines and containing four angles. Polygon – A polygon is a plane figure bounded by more than four straight lines and containing more than four angles. Q43 – What is the difference between a parallelogram and a rhombus? Ans43 – Parallelogram – A parallelogram is a quadrilateral in which the opposite sides are equal and parallel.Rhombus – A rhombus is a quadrilateral in which all the sides are equal and the angles are not right angles. However, in this case the opposite angles ar e equal. Q44 – What is the difference between regular and irregular polygons? Ans44 – Regular polygon – A regular polygon is a plane figure in which all the sides and angles are equal. Irregular polygon – An irregular polygon is a plane figure in which all the sides and angles are not equal. Q45– Name the principal planes of projections. Ans45:-There is two planes employed for projection and are known as reference planes or principle planes of projections.These planes intersect at right angles to each other. These are 1) Vertical plane: – The plane which is vertical is called vertical plane and is denoted by V. P. Vertical plane is also known as Frontal Plane as front view is projected on this plane. 2) Horizontal plane:-The plane which is horizontal and at right angle to the V. P is called Horizontal Plane and it is denoted by H. P. Q46:- What is the principle of projection? Ans46:-If straight lines are drawn from various points on the cont ours of an object to meet a plane, the object is said to be projected on that plane.The figure formed by joining in correct sequence the points at which these lines meet the planes is called the projection of the object. Q47 – What is ground line (G. L. ) or intersection or reference line? Ans47:-The line of intersection of two principle planes of projections i. e. VP and HP is called reference or intersection or ground line and is denoted by x-y line. Q48 – What is an auxiliary view? Ans48:-The view obtained on the auxiliary plane which is parallel to the inclined surface of an object is called auxiliary view. Q49 – What do you understand by missing lines?Ans49 – The lines which are added in the given orthographic projection in order to complete the drawing of an object are called missing lines. Q50 – What do you understand by missing views? Ans50 – The view which is added in the given orthographic projections in order to complete the drawi ng of an object is called missing views. Q51 – What is a sectional view? Why sectional views are used in drawing? Ans51 – The view obtained after cutting the object in order to show the inner details by an imaginary cutting plane is known as sectional view.Sectional views are used in drawing to show the interior details of the object, which are not visible to the observer from outside. Q52 – What is a cutting plane or section plane? Ans52:- The imaginary plane by which the object is assumed to be cut is called the cutting plane or sectional plane. They may be perpendicular or parallel to one of the principle planes and either perpendicular or inclined to the other plane. These planes are represented by their traces. Q53 – What are section or hatching lines? Ans53 – The lines used to represent the material which has been cut by the cutting plane are called section lines.They are also called hatchings or crosshatchings. These are equally spaced lines inclined at 45? to the horizontal. Q54 – What do you mean by sections of solids? Ans54 – the solids which are cut by the section planes to visualize the internal constructional details of the invisible features are known as section of solids. Q55 – What is apparent section? Ans55:- The projection of the section on the plane to which it is inclined is called as apparent section. Q56 – What is true section? Ans56:- The projection of the section on a plane parallel to the plane will show the true shape of the section.Q57 – How will you classify sections of solids? Or What are the different positions of a section plane w. r. t. two reference lines? Or What are the types of sections of solids? Ans57:- 1) Section of solids obtained by horizontal planes. 2) Section of solids obtained by vertical planes. 3) Section of solids obtained by auxiliary inclined planes. 4) Section of solids obtained by auxiliary vertical planes. 5) Section of solids obtained by profile plane. Q58 – What do you understand by V. T. and H. T. of section plane? Ans58 – Horizontal trace (H. T) – H. T. of a section plane is a line in which the plane meets the H.P. Vertical trace (V. T. ) – V. T. of a section plane is a line in which the plane meets the V. P. Q59 – What do you mean by Frustum? Ans59 – When the section plane is parallel to the base plane of a cone or pyramid, it will form a frustum. Q60 – What do you mean by truncated? Ans60 – When the section plane is inclined to the base plane of a solid, it will form a truncated. Q61 – What do you understand by intersection of surfaces? Ans61 – The lines or curves which are formed when surfaces of two solids intersect with each other are known as intersection of surfaces or interpenetration of solids.Q62 – What are the lines or curve of intersection or interpenetration? Ans62:- When a solid penetrates into another solid, their surfaces meet in a line called the line or curve of intersection or interpenetration. Q63 – Give the practical applications of the intersection of surfaces or interpenetration of solids. Ans63:- It is used in (i) sheet metal shop (ii) pipe fittings (iii) boiler fittings (iv) aeroplane construction (e. g. wings, fuse lags etc. ) (v) Automobile layout works (e. g. body wheel house, chairs etc. Q64 – Name the methods of plotting the lines of intersection or inter-penetration of solids? Ans64:- 1) Line method or piercing point method 2) Cutting plane method Q65:- How will you classify the intersecting surfaces? Ans65:-1) the intersection of plane surfaces 2) The intersection of two curved surfaces 3) The intersection of a plane surface and a curved surface Q66 – What do you mean by development of surfaces? Ans66:- A layout of the complete surface of a three dimensional object on a plane surface is called its development or pattern. Q67:- What is stretch out or girth line?An s67:- The stretch out or girth line is the length of the pattern or development and is given by the perimeter of the object measured in a plane at right angles to the axis. This term is used in patterns of objects having a constant cross section for their full length. e. g. prisms and cylinders. Q68 – What is the principle of development? Ans68 – The development is based on the principle which indicates that every line on the development must show the true length of the corresponding line on the surface of the object for which development is required. Q69 – What are the different methods of development of surfaces?Ans69:- 1) Parallel line development 2) Radial line development 3) Triangulation development 4) Approximate method Q70 – Why the true lengths of slant edges are determined? Ans70 – The true length of slant edges are determined because every line on the development must show the true length of the corresponding line on the surface of the o bject to be developed. Q71 – What are the applications of development of surfaces? Ans71:- It is used in the fabrication of simple to highly complicated shapes from flat surfaces in sheet metal shops, in the construction of boilers, pattern making, tunnels, buckets, chimney etc.Q72 – What is a point? Ans72 – A point is that which has simply position but no magnitude. It is generally represented by a very small circle or dot. Q73 – What do you mean by octants? Ans73 – When the three planes i. e. H. P. , V. P. and P. P. divide the entire space into eight quadrants, then these quadrants are known as octants. Q74 – What is the difference between first angle and third angle projection? Which angle projection is recommended by B. I. S. now a days? Or What are the types of orthographic projections? Ans74:-First angle projection:-In this projection the object is assumed to be ituated in first quadrant, i. e. in front of V. P and above HP the project ions obtained on these planes is called first angle projection. The symbol for the first angle projection is Figure. Third angle projection: – In this Projection the object is assumed to be situated in the third quadrant that is below HP and behind VP . The front view comes below the XY line and the top view above it. The symbol for the third angle projection is Now a day we are working with first angle projection because it is recommended by the B. I. S and it is adopted by almost all the countries of the world since 1983.Figure. Q75 – Why the projections of an object is not drawn in second and fourth quadrants? Ans75 – The projections of an object is not drawn in second and fourth quadrants because the overlapping will take place. It will become very difficult to understand the views. Q76 – When the auxiliary planes are used? Ans76 – The auxiliary planes are used in order to view the true shape of an inclined surface. The projection drawn on the auxiliary plane is known as the auxiliary view and gives the true shape of the inclined surface. Q77 – What are the types of auxiliary planes?Ans77:-The plane placed at any angles to the principle planes is called auxiliary plane. Auxiliary planes are of two types. 1) Auxiliary vertical plane (A. V. P. ):-It is perpendicular to the HP and inclined to the VP. Projection on an AVP is called auxiliary front view. 2) Auxiliary inclined plane (A. I. P. ):-It is perpendicular to the VP and inclined to the HP. Projection on AIP is called auxiliary top view. Q78 – Define a straight line. Ans78 – A straight line is defined as the shortest distance between the two points. Q79:- What is true length of a line? Ans79:-When a straight line is inclined to one plane and parallel to the ther, its projections on the plane to which it is parallel will show its true length. Q80 – What do you mean by projections of a straight line? Ans80:-To draw the front view, top view and side view of a straight line is called projection of a straight line. Q81:- What is inclination of a straight line? Ans81:-It is defined as the angle which the line makes with the plane. As such a line has two inclinations i. e. inclination with the HP is represented by an angle and inclination of a line with VP is represented by an angle . Q82 – What are the apparent angles of inclinations?Ans82 – The angle made by the front view of a line with reference line (x-y line) is called apparent angle of inclination ?. The angle made by the top view of a line with reference line (x-y line) is called apparent angle of inclination ?. Q83 – Name the methods to determine the true length and true inclinations of a straight line. Ans83:-The following methods are used when the line is inclined to both the reference planes. 1) Rotation method 2) Auxiliary plane method 3) Trapezoid method. Q84 – What are skew lines? Ans84:-Any two lines that are not parallel with each other and do not intersect are called skew lines.Q85 – What is the trace of a straight line? Ans85:-When a straight line is inclined to a plane, it will meet that plane, produced if necessary. The point in which the line or line produced meets the plane is called its trace. 1) Horizontal trace:-The point of intersection of the line with the HP is called the horizontal trace. 2) Vertical trace:-The point of intersection of the line with the VP is called the vertical trace. Q86 – Define a plane. Ans86:-A flat surface generated by moving a straight line in space is called a plane. A plane fig. has only two dimensions i. e. length and breadth.Q87 – What is the difference between a plane and a lamina? Ans87:-Plane:-A plane has no boundary and it extends to infinity in all directions. Lamina:-The plane which has limited extent is also known as lamina. Q88 – What are the types of planes? Ans88:-There are two types of planes. 1) Perpendicular planes:-The planes w hich are perpendicular to one or both the reference i. e. VP and HP are called perpendicular planes. 2) Oblique planes:-The planes which are inclined to both the reference planes i. e. VP and HP are called oblique planes. Q89 – What is the trace of a plane?Ans89:-The lines in which the planes meet the reference planes i. e. HP and VP are called the traces of the planes. There are two types of traces of planes. 1) Horizontal trace:-The intersection of a plane with the horizontal plane is called the horizontal trace. 2) Vertical trace:-The intersection of a plane with the vertical plane is called the vertical trace. Q90 – What is a solid? Ans90 – An object having three dimensions i. e. length, breadth and height is called a solid. E. g. Prisms, Pyramids, cone, cylinder etc. Q91 – What are different types of solids? Ans91:- Solids may be divided into two main groups. ) Polyhedra or polyhedron: – A polyhedra is defined as a solid bounded by planes call ed faces. Which meet in straight lines called edges? 2) Solids of revolution: – The solids which are formed by the revolution of plane figures are known as solids of revolution. e. g. Cylinders, cones, sphere etc. Q92:- What are right solids? Ans92:- A solid is said to be a right solid if its axis is perpendicular to its base or its end faces. Q93 – What are oblique solids? Ans93:- If the axis of a solid is inclined at an angle other than 90? to its base or end faces, it is called as an oblique solid. Q94:- What are regular solids?Ans94:- If all the edges of the base or the end faces of a solid are equal in length and form regular plane figures, it is said to be a regular solid. Q95 – What is the difference between prism and pyramid? Ans95:- 1) Prism:- A prism is a polygon having two equal and similar end faces, called bases, parallel to each other and joined by other side faces which are rectangles or parallelograms. 2) Pyramid: – A pyramid is a polyhedr on, having a polygon as its base and a number of triangular faces, equal to the number of sides of the base polygon, meeting at a common point called the apex or vertex.Q96 – What are the various positions which a solid can take w. r. t. the reference planes? Ans96 – The following are the different positions which a solid can take w. r. t. the reference planes. (i) The solid resting on base on H. P. , with its axis perpendicular to H. P. , and parallel to V. P. (ii) The solid resting on face on H. P. , with its axis perpendicular to V. P. , and parallel to H. P. (iii) The solids resting on face on H. P. , with its axis parallel to H. P. and V. P. (iv) The solid with its axis inclined to one plane and parallel to the other. v) The solid with its axis inclined to both the reference planes i. e. , H. P. and V. P. Q97:-What is an isometric view? Ans97:- If the projection of an object is so drawn that all the three axis of the object are equally inclined to the plane of pro jection then it is called an isometric view. Q98:- What is an isometric scale? Ans98:- The proportion by which the actual length is converted to isometric length is called as isometric scale. Q99 – What are isometric axis? Ans99 – The three lines OA, OB and OC meeting at a point and making 120? ngles with each other are termed as isometric axis. Q100:- What are isometric and non isometric lines? Ans100:- The lines which are parallel to isometric axis are called as isometric lines. The lines which are not parallel to isometric axis are called non isometric lines. Q101 – What are iso-metric planes? Ans101 – The planes representing the faces of an isometric view of the cube as well as the other planes parallel to these planes are called isometric planes. Q102 – Give the various positions of isometric axis. Ans102 – The various positions of isometric axis are as follows. Figure.

Film Noir to Neo Noir

Murphy 1 Rachel Murphy Professor Charlotte E. Howell Film 2700 12 November 2012 Word Count: 1411 Film Noir to Neo-Noir: A Shift in Cultural Tides Film noir of the 1940s captivated audiences through its distinct form of storytelling. Strongly influenced by German Expressionism, these films have a definitive look and style that still resonates with modern audiences today. Like other classical Hollywood genres, film noir sought to bring to light tensions felt within society, namely those that affected men following World War II.Neo-noir films pay a great deal less attention to social commentary. Like film noir of the past, neo-noir elevates style over narrative; however, the genre has seen significant changes in regards to narrative, the disappearance of the femme fatale, and the prevalence of onscreen violence due to shifting cultural tides. In observing examples of film noir and its contemporary version, neo-noir, it is clear several elements in regards to the style and overall  "feel† of these films have virtually remained the same throughout the years.In Nicolas Winding Refn’s neo-noir, Drive, a sense of otherworldliness is portrayed through several night scenes, intense shadows, and an overall dark rather downtrodden mood to the film. The scenes in the film take place at night and invariably in an urban setting. All of these elements are Murphy 2 extremely typical of classic film noir as well as German Expressionism. Drive’s narrative unfolds with surprisingly little dialogue. Instead Refn focused scenes on the mood, further strengthening the style of the film. Similarly, Curtis Hanson’s L. A.Confidential keeps with traditional film noir in elevating the style of the movie above its narrative. This is done through the heavy emphasis of the urban cityscape. As the title suggests, Los Angeles, is a major component within the film. The peppy, orange-filled paradise portrayal of L. A. in the film’s opening scene sharply cont rasts the corrupt, crime-ridden town shown throughout the rest of the film. In addition, voice-overs and flashbacks, typical elements of film noir, are extensively used. The genre has seen great changes in regards to its social commentary, however.Noir films of the 1940s strongly reflected the social climate of the time. In several respects, film noir can be seen as the male equivalent to melodrama. Just as women dealt with the crisis of femininity in post-war years, men also struggled with their masculinity as well as adjusting to their new roles in an ever-changing society. After World War II, many Americans, especially men who had experienced the atrocities of war firsthand, took on a more cynical outlook on the world. Film noir of the 1940s sought to bring these feelings of isolation and changing attitudes to light.Like many men returning from the war, the heroes were disenchanted and often very isolated. In many respects, their fate is predetermined. In Tay Garnett’s The Postman Always Rings Twice, the audience gains a sense that John Garfield’s character, Frank’s, fate is already sealed as soon as he first plots, and eventually carries out the murder of Cora’s husband. This action clearly serves as a marker in the downward spiral of Frank’s life. Similarly, in Billy Wilder’s Murphy 3 Double Indemnity, Fred MacMurray’s character, Walter, irrevocably alters the course of his life when he gives in to Phyllis’s pleas to murder her husband.In both of these instances, the motivation behind this clearly immoral acts is lust. Both protagonists seem somewhat helpless against these forces. Both films also end with little doubt as to the fate of the protagonists. In The Postman Always Rings Twice, the film ends with Frank awaiting his punishment on death row. Similarly, Wilder’s Double Indemnity ends with Walter, critically injured from a gunshot wound inflicted by Phyllis, confessing his role in her hu sband’s murder. This clearly reflects upon the attitudes of males during the 1940s as helpless against the imposing forces of an oppressive society.Neo-noir films differ from their film noir counterparts because they are no longer reflective on social and cultural tensions. This is simply because the tension is not as widespread or heavily felt in today’s society. In the ending of Refn’s Drive, the nameless driver, though stabbed in the abdomen, clearly lives. It left up to the viewer to decide what kind of life he will lead in the future. In Hanson’s L. A. Confidential, the future of the city is somewhat unclear, but both protagonists in the film are met with at least somewhat happy endings.The male protagonists in neo-noir films are also much more strong-willed. Their actions, though at times extreme, are seen as justified to the viewer and made by the protagonist alone. Unlike earlier noir films, the protagonists are at least somewhat in control of the ir future. This turn within the genre clearly reflects changing attitudes within society, as the helplessness and isolation men felt after the war is no longer felt on such a large scale. Murphy 4 The influence of culture on the content of noir films is especially evident in the disappearance of femme fatale in neo-noir films.The 1940s marked a major shift in gender roles with the start of World War II. As men left for war, women took up jobs in the workforce and in factories in order to help with the war effort. This brought about a new sense of independence for women. When men returned home from the war, however, this shift was not necessarily seen in a positive light. The emergence of the femme fatale in film noir clearly reflects that in the eyes of men, women’s changing roles in society often presented a threat to perceived masculinity as well as established gender roles of the day.The femme fatale of noir films is invariably portrayed in a negative light. She is in most cases seen as the major driving force behind the protagonist’s tragic end. Furthermore, the protagonist is usually helpless against the advances of these women. Femme fatales, such as Cora in The Postman Always Rings Twice and Phyllis in Double Indemnity, are almost always met with an end even more bleak than that of the protagonist. In these two films, the femme fatales are both killed with little thought. Neo-noir films, however, approach female characters in a much more favorable light.The relationships between protagonists and these women are based on love, rather than mere lust. Thus, the actions of the protagonists appear often more justified. This can be accredited to the changing cultural tides since the 1940s. Women’s independence is generally no longer seen as a threat to male masculinity and thus is virtually extinct thematically in neo noir films. This is especially evident in Drive as well. The nameless driver’s love interest, Irene, is characteriz ed by her innocence rather than her sexuality. Murphy 5 Even in L. A.Confidential, Lynn, a prostitute, has a relationship with one of the protagonists, however, the relationship is based on love rather than lust. Film noir arguably would not translate well to modern audiences if not for its integration of onscreen violence. Like German Expressionism, 1940s film noir drew a definitive reaction of discomfort and psychological unease from its audiences. In Double Indemnity, the scene in which Phyllis’s husband is murdered is brief and little is shown. The audience is shown only Phyllis’s cold, detached expression while her husband is murdered next to her in the passenger seat.In the 1940s, filmmakers didn’t necessarily need to show Phyllis’s husband being murdered in order to elicit a strong psychological reaction from audiences. With the abrogation of the Hay’s Code, however, audiences have become somewhat desensitized to the mere implication of viol ence. L. A. Confidential and Drive both use violence as a means of eliciting this same reaction. Perhaps the most memorable scene in Drive occurs in an elevator where the driver, in order to protect himself and Irene, not only kills a man, but proceeds to unleash all of his anger by stomping the man’s head into a gruesome, bloody pulp.In L. A. Confidential, numerous murder scenes and uncomfortable police interrogations illustrate how violence is now used in neo noir to elicit the strong emotional and psychological discomfort that typified 1940s noir. Certainly the strongest influence on the evolution of film noir has been societal and cultural changes throughout time. These changes have served, however, to maintain film noir’s relevance with contemporary audiences while still keeping with specific attention to the overall â€Å"feel† of the film and high level of stylization.

Critical Period and Language Acquisition Essay

Part of the reason why Genie’s case fascinated psychologists and linguists so deeply was that it presented a unique opportunity to study a hotly contested debate about language development. Nativists believe that the capacity for language is innate, while empiricists suggest that it is environmental variables that play a key role. Essentially, it boils down to the age-old nature versus nurture debate. Do genetics or environment play a greater role in the development of language? Nativist Noam Chomsky suggested that the acquisition of language could not be fully explained by learning alone. Instead, he proposed that children are born with a language acquisition device (LAD), an innate ability to understand the principles of language. Once exposed to language, the LAD allows children to learn the language at a remarkable pace. Linguist Eric Lenneberg suggests that like many other human behaviors, the ability to acquire language is subject to what are known as critical periods. A critical period is a limited span of time during which an organism is sensitive to external stimuli and capable of acquiring certain skills. According to Lenneberg, the critical period for language acquisition lasts until around age 12. After the onset of puberty, he argued, the organization of the brain becomes set and no longer able to learn and utilize language in a fully functional manner. Genie’s case presented researchers with a unique opportunity. If given an enriched learning environment, could she overcome her deprived childhood and learn language even though she had missed the critical period? If she could, it would suggest that the critical period hypothesis of language development was wrong. If she could not, it would indicate that Lenneberg’s theory was correct. Genie’s Language Progress Despite scoring at the level of a one-year-old upon her initial assessment, Genie quickly began adding new words to her vocabulary. She started by learning single words and eventually began putting two words together much the way young children do. Curtiss began to feel that Genie would be fully capable of acquiring language. After a year of treatment, she even started putting three words together occasionally. In children going through normal language development, this stage is followed by what is known as a language explosion. Children rapidly acquire new words and begin putting them together in novel ways. Unfortunately, this never happened for Genie. Her language abilities remained stuck at this stage and she appeared unable to apply grammatical rules and use language in a meaningful way. At this point, her progress leveled off and her acquisition of new language halted. While Genie was able to learn some language after puberty, her inability to use grammar (which Chomsky suggests is what separates human language from animal communication) offers evidence for the critical period hypothesis. Of course, Genie’s case is not so simple. Not only did she miss the critical period for learning language, she was also horrifically abused. She was malnourished and deprived of cognitive stimulation for most of her childhood. Researchers were also never able to fully determine if Genie suffered from pre-existing cognitive deficits. As an infant, a pediatrician had identified her as having some type of mental delay. So researchers were left to wonder whether Genie had suffered from cognitive deficits caused by her years of abuse or if she had been born with some degree of mental retardation.

Mile Durkheim Essay Example | Topics and Well Written Essays - 250 words - 1

Mile Durkheim - Essay Example Durkheim’s biggest goal with sociology was that he wanted it to be a quantitatively and scientifically method heavy field. As a result, he wanted to take the scientific method that was used in the natural sciences and use it on society in an effort to describe and predict collective behavior. He believed that a collective consciousness was the glue, which bound everyone together in society. This consisted of beliefs, values, traditions, etc. which all served an important role in unifying society. With the rise of the industrial era and an increase in the segmentation of jobs, he worried that society was changing as the economy was destroying the collective unconsciousness. Instead, it was being replaced upon the reliability and interconnectedness of economic principles. Being one of sociology’s principle founders, Durkheim played an important role in establishing sociology as a scientific field, rather than just as an abstract humanitarian course. This was achieved because of the emphasis he placed on taking the scientific method and applying it to society so that the research and discipline in the field would proliferate. He died on November 15,

Outline of research paper Impact of the Internet on a Small Business Assignment

Outline of research paper Impact of the Internet on a Small Business - Assignment Example Poon (1997) also discussed some research findings regarding competitive advantage and the extent to which the Internet helps in achieving that advantage. The Net Imperative (2009) and other Internet sources were also used as references in making their findings enhanced the contents of this paper. The rest of the paper includes the perspectives and interpretation of the researcher based on the sources used, both offline and online. Among the major findings of the research suggest that the positive impacts outweigh those of the negative. According to Poon (1997), the Internet affects small businesses especially inter-organizational and enterprise-to-consumer because IT costs are relatively low and efficient. Furthermore, the Internet also intensifies relations with trading partners by requesting quotes or perishable stocks of goods through online auction (The Net Imperative, 1999). In addition, the Internet is also a major help in locating and purchasing resources from distant market while it lessens the costs of marketing because it lessens the overseas taxes. In other cases, off-shoring is also a positive impact of the Internet because small companies with high operating cost in their home country can venture into overseas business in countries with lower operational costs. On the contrary, the research also reported the negative impact of Internet in terms of the company’s competitive advantage over b ig industries since both engage in Internet marketing and operations, as well. Lastly, Internet security is also a negative impact since information about the company, as well as its transactions and strategies, are also available via websites and blogs. The Internet has both positive and negative impacts to small businesses. The positive impacts outweigh the negative. Therefore, the Internet is a vital tool in today’s

Final Class Project Case Study Example | Topics and Well Written Essays - 1750 words

Final Class Project - Case Study Example To add on this, it is through documentation that patient response is reflected. On the contrary, the current case lacks documentation from the patient nurse. Here, the nurse testifies that she only had handwritten notes. Furthermore, in her testimony, the nurse says that she did not document any vital signs during the 2 hours of being informed by the patient’s wife and the time the patient is pronounced dead (Guido, 2009). Evidently, the case lacks documentation and this is likely to affect the final outcome. 2. Was negligence there on the part of the nursing staff in the care of this patient? In the case under study, it is evident that the nursing staff showed negligence in the course of duty. The patient nurse is negligent due to failure to document information on the patient. In her evidence, she testifies that she did not have any documented records on vital signs between the time of being informed and the time the patient is pronounced dead (Guido, 2009). As such, it is c lear that no information was available on what went on during the 2 hour period. Additionally, the nurse failed to assess and monitor the patient’s situation after she informed the physician that the patient was bleeding. Given that the patient’s health status could change gradually or suddenly, it was vital for the patient nurse to keep monitoring the situation. On the other hand, the office nurse failed to act as a patient advocate. Here, it is evident that this nurse did not follow up the case to see if the surgeon responded after she informed him. More so, the nurse does not call to check the patient’s progress (Guido, 2009). She only passes the information to the surgeon and that’s it as far as she is concerned. Consequently, it is evident that the nursing department displays negligence while on duty. 3. How does the obvious contradiction in the testimony between the patient’s hospital nurse and the office nurse and the physician’s acco unt of what happened affect your decision in this case? From the case, there is obvious contradiction between the two nurses’ testimony and the physician’s account. This shows a failure to communicate accurately while on duty. There seems to have been lack of communication between the three and hence all are liable to being sentenced for negligence while on duty. Normally, different nursing departments ought to be in constant communication to find the best way possible of helping the patient. This is not the case here. The patient nurse does not keep in touch with the physician to update him on the patient’s situation. Whereas the patient nurse testifies of having informed the physician of the patient’s condition on time, it takes two hours before the physician responds (Guido, 2009). Essentially, the contradiction indicates failure to communicate by all parties involved. 4. What standards for documentation did the patient’s nurse breach? Standards for documentation in nursing entail provision of patient information for communication purposes. Nursing documentation, as a valuable tool, supports effective communication on the patient’s progress for providers. In this particular case, the patient nurse breached these standards. The nurse fails to document the patient’s progress (Guido, 2009). Furthermore, nurses are responsible for their own actions while on duty. As such, documentation forms part of the responsibility. On the contrary, lack of documentation in the case means that the patient’

3rd Report Assignment Example | Topics and Well Written Essays - 1250 words

3rd Report - Assignment Example It is for this reason, among others, that journalists are victimized, harassed, assaulted, imprisoned, and even killed. The categories of journalists who are greatly affected are foreign journalist. This essay will highlight these challenges, by giving an investigative report of two jailed foreign journalists, Laura Ling and Euna Lee. It will examine their work prior to their arrest, their arrest and trial, their sentencing, and the interventions that led to their release. Finally, it will give a conclusion. Euna Lee and Laura Ling are journalists living and working in the United States. Euna Lee is an American of Korean descent while Laura Ling is a Chinese American. They are both journalists at a Television station, Current TV, based in California. Current TV, which broadcasts from San Francisco was co-founded Al Gore who is a former vice-president of the US. Ling works as the station’s news editor, while Lee is the news editor. In the year 2009, the duo began work on a documentary whose intention was to highlight the plight of North Koreans attempting to run away from the dictator government in Pyongyang, North Korea’s capital, into China across the Tumen River (Human Rights Watch, 69). This was the genesis of their troubles with the isolated communist country. The investigative work of Lee and Ling took the duo to China, a longtime ally and cross border neighbor of North Korea but whose laws were a bit friendly to foreign journalists. China was thus a strategic country to launch investigations from. This is mostly so because it shares a border with North Korea which refugees use to cross over. Lee and Ling would thus be able to launch their investigations about human trafficking and the issue of refugees, and tell the story to the world through their documentary. However, they had to conduct all their operations within the borders of China. Otherwise, they would be arrested and

How Free Trade Causes Development Research Paper

How Free Trade Causes Development - Research Paper Example drug barons, arms merchants, rackets bosses, Mafiosi, and other profiteers are emerging as the economic and political leaders of the social transformations underway in their respective societies.† (Buchanan, 2000, p.1) One of the criticisms leveled against free trade as it exists today is its affect on workers and consumers. Some believe that under this system, workers become helpless pawns of their capitalist masters, compelled to sell their labor power at sub-optimal costs. The only theoretical alternative they have to evading this exploitation is to become destitute, which is a far greater misery. Multi-national corporations (MNCs), which are the facade of free trade, are perceived as coercing citizens to unwillingly participate in the capitalist market system, while also leaving consumers with no choice but to buy their products. In the book titled Telling the Truth about History, author Joyce Appleby traces how MNCs came to be the dominant institutions of our age. Here, th e author makes some scathing observations about the nature of capitalist enterprise that is the back bone of prevailing free trade systems: â€Å"One of the distinguishing features of a free-enterprise economy is that its coercion is veiled. . . . The fact that people must earn before they can eat is a commonly recognized connection between need and work, but it presents itself as a natural link embedded in the necessity of eating rather than as arising from a particular arrangement for distributing food through market exchanges....† (Joyce as quoted in Levite, 2002, p.32) The free-trade system is also criticized for promoting 'wage-slavery', whereby human beings are reduced to mechanical automatons as they go through the drudgery of work each day. Here too, the slavery is not so much express... This paper stresses that while free trade has led to development in some countries, they have led to economic instability in others. What is most worrisome about free trade in the modern world is the vacuous nature of the term, as it is stripped of its substantive meaning. In other words, where there is conflict between the execution of this system in its ideal form and the consequences for major business corporations, it is always the interests of the latter that is looked after. This is nowhere more clearly visible than in the history of NAFTA. The terminology can be a little deceptive here, for despite claims of being a 'free-trade' agreement, it has many protectionist provisions in it. A brief survey of the effects of NAFTA on the general population reveals that American, Mexican and Canadian elites have seen most of its benefits. This report makes a conclusion that global free trade arrangements have failed to lead to uniform development. While there are obvious success stories like India, China and South East Asian bloc, much of the rest of the world has not benefitted. It is in response to these failures that the global solidarity movement has arisen. Centered on universal human challenges like poverty-reduction, access to basic healthcare, free education for all children, social welfare for the disadvantaged, etc, the global solidarity movement presents an alternative operative framework to the United States led global capitalist project. In a few decades time, it is plausible that this more pragmatic form of social organization might have quelled American hegemony in economic, cultural and political domains and might have eliminated the need for economic globalization.

Science Development and Computers Coursework Example | Topics and Well Written Essays - 1000 words

Science Development and Computers - Coursework Example This paper would also supplement to entailing the ethical reasons behind making use of computers as fast paced humanoids and also shedding some insight on the much growing artificial intelligence in the world today. Conclusively it might be imperative to reason out the differences in the working of a human brain in retrospect to a central processing unit and how both these entities are in effect the working of the same principal thinking. Science developments and computers With the rapid growth in the information technology industry it is an ever so imperative fact that fast and reliable data transfers are the key to enterprise success in the upcoming future. While organizations battle out for the highest gains and profits, all it comes down in the end is to the fact that which company pertains to the fastest solutions and outputs, whereas this being the age of the integrated technological advancements it is no wonder that a lot has been researched for necessitating new methods and w ays of providing fast data transfer and processing techniques. Before lunging into the technical specification of when and how an industry can optimize its information technological feats it is profoundly imperative that we seek the reasoning and logic behind as to what causes an uplift in most technological advancements. ... With time we have seen that those IBM personal computers have been effectively reduced into mere handheld smartphones today having more than four times the amount of memory and processing speed those early vintage personal computers could potentially offer. With this type of an increasing change in trend and usage we can establish the datum that size of the object has significantly affected the experience these computers provide today and with specification from the study of material science it can duly be distinguished that ascertaining to the working size in a technological development makes way to a much compacter solution to the same specifications while sometimes working on molecular level can yield great optimizing results. With such reasoning we can surly address that the three things imperatively necessary to influence any sort of material and integrated source are firstly the kind of atoms that embed the structure for example the use of silicon atoms as opposed to copper one s idealize the functionality of a microchip while subsequently how these particular atoms are bonded and arranged are the preceding qualities that are in close check with computational hardware development. The reactivity, density and malleability are also features that ascertain a physical influencing change but on a more fundamental aspect of modeling hardware the atomic size structure and behavior are the key material things that effectively model a wiser choice of technology. Coming back to the usage of silicon and how fast this tech seems to cater have fundamentally establishes that the developments in Silicon technology and the express

Leases Essay Example | Topics and Well Written Essays - 1250 words

Leases - Essay Example IAS 17 has undergone many revisions in the past to improve its application. However, even with all the amendments and revisions, it still has some issues that cause problems in its application in accounting for leases. The purpose of IAS 17 is to classify leases and provide the policies to guide how a firm treats leases in its financial statements. The standard has a few exceptions concerning some lease agreements. Some assets are also not part of the standard, therefore, their recognition and treatment do not fall within the standard. For the leases and assets that qualify within the standard, classification divides them into finance and operating leases (IFRS, 2013a, p.771). The determination of the class of a lease as either a finance lease or an operating lease is at the commencement of the lease agreement. IAS 17.4 defines the two classes of leases giving their characteristics. The substance of the transaction, as opposed to the form, determines whether a lease is a finance or operating lease (IFRS, 2013a, p.772). The treatment and accounting for the two classes of leases differs. The lessee recognizes operating leases as an expense. The lease payments appear in the income statement as an exp ense over the lease term and follow a straight-line basis. The lessor recognizes assets under lease in the balance sheet. The income statement of the lessor recognizes the lease income on a straight-line basis. As for finance leases, their accounting involves recognition both as an asset and liability in the books of the lessee. The value to appear in the books is either the asset’s fair value or the present value of the minimum lease payments, whichever is lower. The lessee also accounts for the depreciation of the asset in accordance with the existing depreciation policies and apportions lease payments between the reduction of the outstanding liability and the finance charge. On the lessor’s books, the accounting for a

Philosophy of Language Essay Example | Topics and Well Written Essays - 2500 words

Philosophy of Language - Essay Example The group was called logical positivists, and they accomplished this union by introducing the notion of convention. The class of positivists asserted that logical empiricism was possible through convention or arbitrariness, where agreement would be reached on the meaning of statements. Quine sought to attack this school of thought through a two-pronged approach; reliance on reductionism and making a distinction on analytic and synthetic distinctions. Dogmas are sets of beliefs that are held to be true by certain people without question, and are often called such as a mark of disapproval from an observer or analyst. Quine felt that logical empiricism lacked legitimacy because of two dogmas; one of them was the distinction between analytic and synthetic truths. An analytic truth may be understood as an assertion that is true exclusively because of its meaning while a synthetic one is held as such owing to facts. Quine felt that the distinction between analytic and synthetic sentences was baseless by looking at a series of assumptions and definitions in the school of thought. Quine started with the notion of synonyms where logical positivists claim that a sentence may be defined as analytic if synonyms can be used to substitute original words and the expressions remain logical truths (Schwitzgebel, 2008). However, the philosopher opposed this statement because it presupposes that synonymy is a well understood and defined term, yet it needs to be explained before it can be applied. The philosopher sought to look for other ways in which logical empiricists sought to defend themselves, such as by saying that a logical truth exists if a sentence has terms whose definitions can be substituted by others. Quine has a problem with this component as well because it is not clear whether an irregularity can arise because of the meaning of the term or the belief that one holds about it.

Disgusting at the same time Essay Example for Free

Disgusting at the same time Essay The author wants the reader to imagine the most horrible things and as everyone has different ideas about our own horror it will make it even more repulsive. By using the word hollowed he burns an image in your mind and makes you visualize the hollow bodies that had been devoured by the vultures. The word Strange by itself in one line sums up your feelings among the following section of the poem, and by being alone in one line it emphasises the word, it gives the word Strange a lot of importance. Achebe shows affection as a pessimistic aspect of life, in the poem it says that love coils up like a snake in a corner, it also says that love is upset, angry or punished. Together with the phrase turned to the wall, the author personifies love. Reaching to a certain point of the poem, the author uses an ellipsis by dividing it into two supposed different stories, however, thats what it seems from the outside, but if you , both stories is related one to another. To link these parts, the author changes line, and uses punctuation ( ), he uses three dots at the end of the first part to show the poem continues, and then starts talking about the commandant Thus the Commandant at Belsen, which appears to be a total different theme. When the poet uses the phrase fumes of human roast it intends to create a disgusting scene, with the word roast he creates a linking image which relates the phrase to the animals, food and cooking (burning). The word i roast` is associated to the word ihuman` which makes you think of people being cooked and burned, and it seems even more revolting as the reader probably visualises itself in the same situation. With this extremely inhuman scene the author originates a cruel image referred to Commandant, he is also shown as a very horrendous man when Achebe talks about the commandants appearance; i hairy nostrils`, the poet wants to incite the reader to hate this character. The Commandants children are represented as his `tender offspringi , this produces a comparison between the commandant and the vulture because normally when referring to society the `offspringi of someone usually are their sons or daughters, the word `offspringi is applied when we talk about animals, so this word in a way shows that the commandant wasnt very loving towards his children. The word tender is used to describe is normally used to describe soft meat. This creates two impressions of the same concept; his offspring is related to good meat, yet its also related to the vultures, which creates a memorable paradoxical image. The author wants the audience to see both facets of this terrible man, by saying the word Daddys, this makes the commandant seem sweet and caring, and uses an enjambment Daddys // return, to make the word i returni stand-out. He also wants to create two different images with the word return, to make the reader think that the children miss their father, and to prove that theres also a bit of grace in such a cruel man. To conclude, in the last paragraph Achebe summarises the poem. He thanks God that even an ogre (which in society is seen as a stereotype of a malicious creature) has a tiny glow-worm of tenderness encapsulated in icy caverns of a cruel heart. This means that all human kind beings with a dark inside will unfailingly have a spark of mercy in him. Achebe finally expresses that human beings arent good or bad, theyre a combination of both, and this is what the whole poem represents. The poem is made out of one stanza, which is divided into four subsections. This an unusual poem because the poet uses free verse, which makes the poem colloquial. It has no rhyme because rhymes make things amusing and musical and wouldnt help the poet describe pessimistic aspects as he does in the majority of the poem. The four fragments link together evil, goodness, vultures and the commandant. Achebe uses commas and enjambment to make it a slow paced poem to read which makes it sorrowed. The whole poem is written in English by a Nigerian author, it is written for European readers. He wants to show that it doesnt matter from where you belong, every war is the same as abominable and everyone has a bit of light and darkness in their hearts.

Predictors of Patient Satisfaction

Predictors of Patient Satisfaction CHAPTER 1: Introduction In this chapter we will briefly discuss the background of the research area. At first an overview of the patients satisfaction concept will be introduced. This will be followed by the relevance of this thesis to the country of Kuwait. After that, we will present a discussion of the problem that will give a better understanding to the reader for our subject. The chapter ends with a brief description of the thesis structure follow. 1.1 Overview: The patient experience reflects the way in which a person perceives the sum total of his experiences with the health system throughout the continuum of care. This experience is influenced by the total of the encounters between the patient and the caregiver, of the patient’s expectations from the health system and from the organizational culture in all of the frameworks with which the patient comes in contact. In recent years, the patient experience has become a central talking point in the ongoing discourse of the health systems in Israel and worldwide, which coincides with numerous social trends: Increasing public awareness of the patient’s rights. Public demand for transparency in conduct and for exploiting opportunities to improve. Activity of patient safety organizations. Changes in the service and consumer cultures. Demand for equality and accessibility to each and every individual. Increasing use of electronic communication and social media in search of medical information, knowledge sharing among patients, recommendation of specialists, alternative treatments and facilities and so on. Patient satisfaction is generally considered as the extent  to which the patients feel that their needs and  expectations are being met by the services provided[4]. Patient satisfaction predicts both compliances [5] and  utilisation [6] and may even be related to improved  health [7]. It also contributes to the atmosphere  prevailing in a PHCC [7,8]. It is associated with  continuity of care [3], the doctors communication skills  [9], the degree of his or her patient centeredness [10]  and the congruence between intervention desired and  that received by the patient [11] . Other factors  influencing satisfaction with medical care include  confidence in the system and a positive outlook on life  in general[12] . Finally, satisfaction is the judgment of  the patient on the care that has been provided [13]. The  physician remains a key element in patient satisfaction  [14]. Summarize chapter 2 in one a half page start with small problem definition, actually there are determinants 1.2 Relevance of this thesis to the country of Kuwait With the huge growing number of cancer patients worldwide in recent years the needs of effective and capable health care suppliers is mandatory. The health care sector in Kuwait has been occupying very important position among the public, as it takes the responsibility to maintain peoples heath and to prevent disease as well to secure complete coverage of the health care services. Kuwait consists six general hospitals for several specialty, and more than 70 care clinics. With an aging facilities and growing population, hospitals have become overused with empower of tools and equipments and the care clinics are not serving dangerous situations or real disease. Cancer cases in Kuwait has reached the top of disease and have been recognized as a main cause of deaths in the country (Alduaij 2012). Kuwait control cancer centre (KCC) comprehensive cancer center have 600 qualified medical staff and 459 beds. KCCC treats over 2000 new cancer patients each year and total of 28,697 from Kuwait and the region(Kuwaitcancercenter.com 2014)[50]. Identification of predictors of patient satisfaction (what aspects of care matter the most to patients) enables policy makers at the Ministry of Health in Kuwait to focus on these aspects and improve them. The correlates of socio-demographic characteristics of patients with satisfaction allow the health care providers to cater to the different needs of patients based on their socio-demographic characteristics. This study aims at identifying predictors of patient satisfaction in the primary care clinics of the Ministry of Health, Kuwait (factors leading to patient satisfaction or dissatisfaction) and its socio-demographic correlates. 1.3 Problem Definition: Lack of proper strategies and support system by Ministry of Health (MOH). There are many obstacles interfering the treatment of patients in Kuwait due to poor system and insufficient management strategies that is related to health care services (Alduaij 2012). Kuwait Cancer Control Center (KCC) which was built back in the 70s is considered as one of the important medical centers in Kuwait for the reason of increasing number of cancer patients in Kuwait (KCC 2014)[51]. KCC have a capacity of 459 beds against 28,697 patients with 1643 employeed,490 nursed, and 11 physicians. Based on that, the hospital have witnessed a migration of the experienced staff which results in shortage of human resources and accordingly leads to instability and sub-optimal medical system (Annaharkw.com, 2014)[53]. Huge public demand to improve hospital services. According to Arab Times (2014)[52] the previous Kuwaiti parliament members have step against the low performance of Ministry of Health (MOH) and of KCCC in particular. They have raised their disagreement over the government’s failure to control the disease and to provide adequate services to satisfy the patients. Many issues were declared to the ministir of health seeking for urgent solutions such as: long waiting time, unavailability of beds , lack of the hospital human resources and machinery as well financial and administrative capabilities. Temporary recovery plan but not permanent solutions. His Highness the Amir Sheikh Sabah Al-Ahmad Al-Jaber Al-Sabah, at the end of year 2013 gave instructions to the government to send cancer patients for treatment abroad at the States expense and to expedite with actions (Kuna, 2014)[50]. The reason for the urgency of this new law was due to public dissatisfaction, and the continuous complaints toward cancer treatment in Kuwait. Dire personnel experience A personnel experience with the cancer disease has been the real motivation behind this study. Watching a dear person suffering not just because of the disease, but because of shortage of resources and lack of proper support system, is the hardest thing ever especially when it comes to losing that dear person forever. 3.6 Research objectives The research objectives are as the following: To explore and review the available international literature about the cancer patents satisfaction. To discuss the determinant of cancer patients satisfaction. To identify service quality dimensions related to cancer patients. To investigate the importance of quality of life for cancer patients. To find out the impact of socio demographic characteristic on cancer patients in Kuwait. 3.7 Research questions Based on the objectives of the study, key questions should be addressed: 1. What are the determinants needed to reach an effective cancer patients satisfaction in Kuwait? 2. What are the health related service quality dimensions? 3. What are the quality of life factors that contribute cancer patients satisfaction? 4. What is the effect of socio-demographic characteristic on cancer disease in Kuwait? 1.6 Research Methodology Our research is considered deductive, quantitative, descriptive and explanatory based on the experiments presented in our literature review. This study is focusing on cancer patients and the benefits that are provided by KCCC hospitals and their personnel life. The questionnaire is in two language English and Arabic and have been randomly distributed in Kuwait cancer control centre and on website. The study population consisted of the patients who came for therapy in KCCC ( outpatients) and the sample size consisted of 300 patients based on total cancer patients in Kuwait which are approximately 28,697 patients. The eligibility criteria included patients who have been diagnosed with different types of cancer as a minimum of 6 months ,above 21 years old and currently are undergoing treatment. The questionnaire contained a socio-demographic characteristics as well as the overall satisfaction with the different aspects of quality of services and quality of life. At last, The data analys is is obtained through using different statistical techniques by using the SPSS software version 17. 1.7 Thesis structure This thesis is divided into five chapters. In the first chapter, we will be providing a background and an overview of the selected research subject, followed by the problem area discussion and description of the thesis structure. In chapter two, an academic literature review related to patients satisfaction and the different independents variables such as physcian concern,staff concern, convenience of care process, tangibles, social well-being, emotional well-being, and information knowledge will be introduced based on theories, academic studies and reports. Chapter three explains the methodology and techniques that have been used in our thesis to analyze and carry out this study. In chapter four, an analysis of the empirical data will be presented along with a discussion of the survey findings and results. Last chapter, number 5, is summarizing the results that have been achieved in our study along with a comparison with other studies. At the end of the final chapter, we will be pro viding a conclusion and recommendations for management and future research. A definition of quality of life The quality of life can only be described and measured in individual terms, and depends on present lifestyle, past experience, hopes for the future, dreams and ambitions. Quality of life must include all areas of life and experience and take into account the impact of illness and treatment. A good quality of life can be said to be present when the hopes of an individual are A good quality of life is therefore usually expressed in terms of satisfaction, contentment, happiness and fulfillment and the ability to cope. This definition emphasizes the importance of personal growth. (K. Chambers et al., 2011) Calman, K. 1984. Quality of life in cancer patients -an hypothesis. Journal of medical ethics, [Accessed: 16 Nov 2013].

Sperm Assessment Using Flow Cytometry

Sperm Assessment Using Flow Cytometry State of the art in sperm assessment using flow cytometry Abstract Flow cytometry is emerging as an important tool in the field of modern andrology for routine analysis of spermatozoa. Recently, application of flow cytometry in the artificial insemination industry especially for pig is a new approach. Until very recent, semen sample analysis was routinely performed by microscopical evaluation and manual techniques by laboratory operators; the analysis is affected by a wide imprecision related to variability among observers, influencing its clinical validity. The last decade, several new flow cytometric techniques have been introduced for farm animal semen assessment that enable a more detailed evaluation of several sperm characteristics. Here in this paper, an initiative has been taken to focus on a number of recent flow cytometry developments important for addressing questions in andrological tests. After the invention of flow cytometry, sperm evaluation by traditional microscopic means became questioned due to the robust advantages of flow cytometry over the microscopic method. Due to the recent development of large number of fluroscence probes, flow cytometry is now capable of analyzing number of sperm characteristics like viability, capacitation, acrosomal integrity, membrane permeability, membrane integrity, mitochondrial status, DNA integrity, decondensation of DNA and differences between gamets based on sex. The application of flow cytometry to their detection allows increased numbers of spermatozoa to be assessed over a short time-period, provides the possibility of working with small sample sizes, increases the repeatability of assessment, removes the subjectivity of assessment and allows simultaneous assessment of multiple fluorochromes. Flow cytometry is a technique capable of generating significantly novel data and allows the design and execution of experiments that a re not possible with any other technique. Nowadays, semen evaluation using laboratory assays is extremely important to the artificial insemination industry to provide the most desired quality product to customers. Future development of flow cytometric techniques will permit further advances both in our knowledge and in the improvement of assisted reproduction techniques. In this paper, the main semen parameters that can be analyzed with fluorochromes and adapted for use with a flow cytometer will be reviewed and the relationship of these tests to fertility will be discussed. Introduction Semen evaluation is the single most important laboratory test that has helped us to identify clear-cut cases of fertility (Jarow et al., 2002), infertility or even of potential sub-fertility (Rodrà ­guez-Martà ­nez, 2007). Determination of the potential fertility of semen sample and, in the long run, of the male from which it has been collected is the ultimate goal of semen evaluations in clinically healthy sires. Methods are available that can sometimes estimate the potential fertilizing capacity of a semen sample and, in some cases, of the male (reviewed by Dziuk 1996; Rodrà ­guez-Martà ­nez et al. 1997a; Rodrà ­guez-Martà ­nez and Larsson 1998; Saacke et al. 1998; Larsson and Rodrà ­guez-Martà ­nez 2000; Rodrà ­guez- Martà ­nez 2000, 2003; Popwell and Flowers 2004; Graham and Mocà © 2005; Gillan et al. 2005). The methods routinely used for evaluation of the quality of a semen sample involved an evaluation of general appearance (i.e. colour, contamination, etc.), volume, pH, sperm concentration, viability, morphology and motility. Most of these techniques are microscopic analyses that only measure a small number of spermatozoa within a population, are time-consuming, can be subjective and generally measure sperm attributes individually. Recently, limitations of semen evaluation methodology have been brought into sharp focus by controversies raised in the epidemiological literature. It should also be noted that such conventional measurements are prone to extreme inter-ejaculate variation, even when the laboratory methodology has been standardized. In the wake of this information, new opportunities have arisen for the development of methods for the diagnosis of male infertility, many of which have been shown to exhibit a prognostic value that eludes conventional semen profiling. Moreover, ejaculated spermatozoa are nowadays handled for use in assisted reproductive technologies, such as the artificial insemination of chilled, frozen-thawed or sexed se men, and IVF. Such handling implies semen extension, fluorophore loading, ultraviolet and laser illumination, high-speed sorting, cooling and cryopreservation, procedures that impose different degrees of change in sperm function following damage to sperm membranes, organelles or the DNA. Therefore, although several assays have been developed to monitor these sperm parameters, recently it is being claimed that buck of these procedures are incomplete, time consuming and laborious. Flow cytometry in different technical applications offers many advantages for the analysis of sperm quality. Flow cytometry allows the simultaneous measurement of multiple fluorescences and light scatter induced by illumination of single cell or microscopic particles in suspension, as they flow very rapidly through a sensing area. The increasing use over the past decade of flow cytometry in the leading laboratories in human and veterinary andrology has dramatically increased our knowledge of sperm function under physiological and biotechnological conditions. Flow cytometers can acquire data on several subpopulations within a sample in a few minutes, making it ideal for assessment of heterogenous populations in semen sample. Initially developed in the 1960s, flow cytometry made automated separation of cells based on the unique recognition of cellular patterns within a population feasible (Hulett et al., 1969). Using such a separation approach, cellular patterns can be identified by as sessing, in individual cells within a population, protein expression using fluorescently labeled antibodies and other fluorescent probes (Baumgarth and Roederer, 2000; Herzenberg et al., 2006). Flow cytometry was first developed for medical and clinical applications such as haematology and oncology. These areas still account for the vast majority of publications on this technique, but during the past few years it has been used in other areas, such as bioprocess monitoring, pharmacology, toxicology, environmental sciences, bacteriology and virology. Recent advancement of flow cytometry increased its application in the reproductive biology especially for andrology. FCM is increasingly used for basic, clinical, biotechnological, and environmental studies of biochemical relevance. Although flow cytometry may overestimate the population of unlabelled cells (Petrunkina and Harrison, 2009), plethora of research from our group in pig (Pena et al., 2003, 2004, 2005; Spjuth et al., 2007; Fernando et al., 2003; Saravia et al.,2005, 2007,2009; De Ambrogi et al., 2006; ) bull (Bergquist et al., 2007; Nagy et al., 2004; Januskauskas et al., 2003; Bergqvist et al., 2007; Hallap et al., 20 05, 2006;) stallion ( Kavak et al., 2003; Morrell et al., 2008) indicate that newly developed fluorescent stains and techniques of flow cytometry has made possible a more widespread analysis of semen quality at a biochemical, ultrastructural and functional level. Therefore, flow cytometry is the current technical solution for rapid, precisely reproducible assessment of sperm suspensions. In this review we have described potentiality and scope of flow cytometry for the evaluation of semen, and the way in which this technique can be used in clinical applications for andrology based on some of our previous experiences. Definition of flow cytometry The definition of a flow cytometer is ‘an instrument which measures the properties of cells in a flowing stream. In other word, a flow cytometer will be defined as ‘an instrument that can measure physical, as well as multi-colour fluorescence properties of cells flowing in a stream. In other work, cytometry refers to the measurement of physical and/or chemical characteristics of cells or, by extension, of other particles. It is a process in which such measurements are made while the cells or particles pass, preferably in single file, through the measuring apparatus in a fluid stream. The data obtained can be used to understand and monitor biological processes and develop new methods and strategies for cell detection and quantification. Compared to other analytical tools, where a single value for each parameter is obtained for the whole population, flow cytometry provides data for every particle detected. As cells differ in their metabolic or physiological states, flow cytometry allows us not only to detect a particular cell type but also to find different subpopulations according to their structural or physiological parameters. Flow cytometry is a technique for measuring components (cells) and the properties of individual cells in liquid suspension. In essence, suspended cells are brought to a detector, one by one, by means of a flow channel. Fluidic devices under laminar flow define the trajectories and velocities that cells traverse across the detector, and fluorescence, absorbance, and light scattering are among the cell properties that can be detected. Flow sorting allows individual cells to be sorted on the basis of their measured properties, and one to three or more global properties of the cell can be measured. Flow cytometers and cell sorters make use of one or more excitation sources and one or two fluorescent dyes to measure and characterize several thousands of cells per second. Flow cytometry gives objective and accurate results (Bunthof et al., 2001; Shleeva et al., 2002), overcoming the problems with the manual methods described above. Function and types of flow cytometry Fluidics, optics and electronics are the three main systems that make up a flow cytometer. In a few minutes, the flow cytometer can acquire data on all subpopulations within a sample, making it ideal for assessment of heterogenous population, such as spermatozoa. The adaptation of flow cytometry to sperm assessment began when it was used for measuring their DNA content (Evenson et al., 1980) and its application to semen analysis has gradually increased over the last 10-15 years. Flow cytometry is now applied to semen evaluation of traits such as cell viability, acrosomal integrity, mitochondrial function, capacitation status, membrane fluidity and DNA status. New fluorescent stains and techniques are continuously being developed that have potential application to the flow cytometric evaluation of spermatozoa. Flow cytometry permits the observation of physical characteristics, such as cell size, shape and internal complexity, and any component or function of the spermatozoon that can be detected by a fluorochrome or fluorescently labeled compound. The analysis is objective, has a high level of experimental repeatability and has the advantage of being able to work with small sample sizes. Flow cytometry also has the capacity to detect labeling by multiple fluorochromes associated with individual spermatozoa, meaning that more than one sperm attribute can be assessed simultaneously. This feature has an added benefit for semen analysis, as few single sperm parameters show significant correlation with fertility in vivo for semen within the acceptable range of normality (Larsson and Rodriguez-Martinez, 2000) and the more sperm parameters that can be tested, the more accurate the fertility prediction becomes (Amman and Hammerstedt, 1993). There are two main types of flow cytometers-analysers and sorters. Sorters have the ability not only to collect data on cells (analyse cells) but also to sort cells with particular properties (defined by the flow cytometer operator) to extremely high purities. There are also a number of commercial flow cytometers that have been developed for particular analytical requirements. Partec manufacture a Ploidy Analyser and also a Cell Counter Analyser. Optoflow has developed a flow cytometer for the rapid detection, characterization and enumeration of microorganisms. Luminex is developing technology for multiplexed analyte quantitation using a combination of microspheres, flow cytometry and high speed digital processing. Advantages of FC compared to other conventional techniques to explore sperm structure and function During the past 2 decades, there has been an increasing interest in reliable assays for assessing semen quality in the fertility clinic and artificial insemination industries. The use of flow cytometry for sperm analysis is an attempt to address the long-standing problem of the subjective nature of the manual method commonly used for semen analysis. An additional source of laboratory variation is the low number of sperms analyzed with manual techniques. Because of time and cost restraints, most laboratories analyze only 50 to 100 sperm to compute the percentage of each cell population and the viability rate. This small sample from a population of millions probably results in a statistical sampling error (Russel and Curtis, 1993). The conventional methods used are limited to microscopic determination of sperm concentration using a hemocytometer (Jorgensen et al., 1997) and evaluation of sperm motility and morphology (Keel et al., 2002). These methods usually involve a subjective asses sment of a few hundred sperm, and quality assurance is rarely implemented in the laboratories performing such analysis. Flow cytometry is a technique that is superior to conventional light microscopy techniques in terms of objectivity, number of cells measured, speed, and precision (Spano and Evenson, 1993). The technique has been used on human sperm to determine a number of factors, including membrane integrity, mitochondrial function, acrosome status, and multiparameter measurement (Garrido et al., 2002). Flow cytometry permitted us to analyze thousands of cells in few seconds. In our series of studies, we demonstrated the feasibility and reproducibility of an automated method to evaluate sperm cell type, count, and viability in human boar samples. In our hand, the precision of the flow cytometric analysis is satisfactory in diverse species (boar, bull, stallion etc), and the observed CVs were significantly better than those reported for the manual method. While there are many advantages of using the flow cytometer for routine semen analysis, its use is often limited to research by the expense and difficulties of operation associated with the requirement of a skilled operator. In addition, a flow cytometer is quite large and cannot withstand shocks associated with movement, meaning it requires a dedicated position in the laboratory. However, the development of more affordable ‘‘bench-top flow cytometers has recently increased the potential application to semen analysis. If we consider flow cytometric analysis further, we can see that it is gaining wider acceptance as a technique for assessing the acrosome reaction and viability simultaneously. Comparing these assays to the more widely used epifluorescent microscopic techniques, the flow cytometric analysis is able to give a far more simple and objective method of analysis, especially with regard to correlation of fertilization with acrosome reactivity potential (Uhler et al., 1993; Purvis et al., 1990; Carver-Ward et al., 1996). A large number of different techniques to estimate sperm concentration have been reported. In the mid-1990s a series of fixed-depth disposable slides were evaluated as rapid and effective pieces of equipment for the estimate of sperm concentration. Preliminary data from a number of studies suggested that, at least in the 20-mm-depth format, such chambers resulted in a noticeable underestimate of sperm concentration compared to the gold standard (improved Neubauer hemocytometer). Using this information, the World Health Organization stated that ‘‘such chambers, whilst convenient in that they can be used without dilution of the specimen, may lack the accuracy and precision of the haemocytometer technique (World Health Organization, 1999). Further data—for example, from Tomlinson and colleagues—showed that 2 proprietary disposable slides (Microcell, Conception Technologies, San Diego, Calif; Leja, Leja Products, BV Nieuw- Vennep, The Netherlands) gave lower spe rm concentrations compared to the hemocytometer method (Tomlinson et al., 2001). To put this in context, numerous reports document unacceptable discrepancies between different laboratories and even between different individuals, although fewer studies attempt to address these issues. So, what is wrong? Several reports emphasize the need for improvement in overall quality of semen testing within and between laboratories (Neuwinger et al., 1990; Jorgensen et al., 1997; Keel et al., 2000). However, the subjective nature of conventional semen analyses, combined with their relatively low precision due to the low number of cells assessed, leads to poor intra- and interlaboratory reproducibility; therefore, the introduction of standardized or quality controlled procedures will probably have a limited effect. The conventional analyses are used to determine whether parameters obtained from an ejaculate are within the range characterized by fertile men, and these methods can therefore provide only unclear cut-off values when used for the prediction of fertility status. Many of the advantages that accrue when using flow cytometry may, when applied to assessment of sperm cells, help overcome some of the mentioned problems found in conventional semen analysis. In the field of semen analysis, validation of a method is important because it is essential to have specific, precise, objective, and accurate laboratory tests to establish a correlation of the data with fertility or to determine the fertility potential of a semen sample correctly (Amann, 1989). Precision of a laboratory test is of great concern to the andrologist in the fertility clinic, since the results of the semen analysis are often used to advise a patient about his fertility and the prognosis for the treatment of the couple. To use established cut-off values and ensure uniform diagnosis, within and between laboratory variations should be determined and followed closely. Accurate determination of sperm cell concentration is critical to the AI industry because it provides assurance both to bull studs and to customers that straws of extended semen contain the sperm numbers indicated. An accurate measure of sperm concentration is particularly important in export markets in which verification of numbers may be required. Routine sperm counts can help to identify possible processing errors within a specific batch of semen or on a particular day, should those errors occur. As sperm counting procedures become more refined, routine counting can be used to monitor subtle changes in daily semen processing that might affect the number of sperm packaged in a straw. Hemacytometers are widely used for routine sperm counts, but the equipment is slow, and multiple measurements of each sample are needed. Single hemacytometer counts are not highly accurate; because of inherent errors in the technique, Freund and Carol (13) found that mean differences of 20% were not uncommon between duplicate sperm count determinations by the same technician. Electronic counters provide much more rapid counting, are easier to use, and give more repeatable results among technicians. However, those instruments tend to include in the sperm count any somatic cells present, immature sperm forms, cytoplasmic droplets, debris, and bacteria, thereby inflating the concentration value (19). Currently, the primary method used by the AI industry to estimate sperm concentration is spectrophotometric determination of turbidity of a semen sample using an instrument previously calibrated for sperm concentration with a hemacytometer or Coulter counter (1). This approach is only as accurate as the methods used for spectrophotometer calibration. New, more accurate methods for sperm count determinations are being sought to replace the older ones. Some laboratories are trying the Maklerm counting chamber (Seif- Medical, Haifa, Israel) and other improved hemacytometers, such as the MicroCellTM (Fertility Technologies, Inc., Natick, MA); however, these techniques will likely have standard lems similar to those associated with the standard hemacytometers. It may be argued that when comparing fluorescent microscopy assays with flow cytometry, one is examining patterns of fluorescence rather than fluorescence intensity, i.e., the flow cytometer is not capable of discriminating sperm which have a fluorescent marker bound to the equatorial segment or over one of the acrosomal membranes (Parinaud et al., 1993; Mortimer and Camenzind, 1989; Mortimer et al., 1987). Tao et al. (1993) compared flow cytometry and epifluorescent microscopy with various lectins and indicated that there is no significant difference between the two methodologies for detection of the acrosome reaction. However, it has been argued that lectins do not bind specifically to the acrosomal region of the sperm (Purvis et al., 1990; Holden and Trounson, 1991) and that other binding sites can be easily distinguished by epifluorescence microscopy, whereas flow cytometry identifies the signal from the entire sperm. Additionally, conventional light microscopic semen assessment is increasingly being replaced by fluorescent staining techniques, computer-assisted sperm analysis (CASA) systems, and flow cytometry (PenËÅ"a et al., 2001; Verstegen et al., 2002). Additional advantages over existing techniques are that this approach is faster than the hemacytometer and that cellular debris, fat droplets, and other particulate material in extended semen are not erroneously counted as sperm, as often occurs with electronic cell counters. This method can also be used to determine the number of somatic cells in a semen sample. Viability The viability of spermatozoa is a key determinant of sperm quality and prerequisite for successful fertilization. Viability of spermatozoa can be assessed by numerous methods, but many are slow and poorly repeatable and subjectively assess only 100 to 200 spermatozoa per ejaculate. Merkies et al. (2000) compared different methods of viability evaluation. They concluded that Eosin-nigrosin overestimate viability while fluorescent microscope and flow cytometry estimate similar trend of viability. Currently flow cytometric procedures have been developed which simultaneously evaluate sperm cell viability, acrosomal integrity and mitochondrial function. This method has been successfully used for assessing spermatozoa viability in men (Garner and Johnson, 1995), bulls (Garner et al., 1994; Thomas et al., 1998), boars (Rodrà ­guez-Martà ­nez, 2007; Garner and Johnson, 1995; Garner et al., 1996), rams (Garner and Johnson, 1995), rabbits (Garner and Johnson, 1995), mice (Garner and Johnson, 1995; Songsasen et al., 1997), poultry and wildfowl (Donoghue et al., 1995; Blanco et al., 2000) and honey bees (Collins and Donoghue, 1999; Collins, 2000) and in fish (Martin Flajshans et al., 2004). Considerable information has accumulated on the use of fluorescent staining protocols for assessing sperm viability (Evenson et al., 1982). The SYBR 14 staining of nucleic acids, especially in the sperm head, was very bright in living sperm. Good agreement was observed between the fluorescent staining method and the standard eosin-nigrosine viability test; the flow cytometric method showed a precision level higher than that of the manual method. One of the first attempts to assess sperm viability utilized rhodamine 123 (R123) to assess mitochondrial membrane potential and ethidium bromide to determine membrane integrity using flow cytometry (Garner et al., 1986). Other combinations that have been used to examine the functional capacity of sperm are carboxyfluorescein diacetate (CFDA) and propidium iodide (PI) (Garner et al., 1988; Watson et al., 1992); carboxydimethylfluorescein diacetate (CMFDA), R123, and PI (Ericsson et al., 1993; Thomas and Garner, 1994); and PI, pisum sativum agglutinin (PSA), and R123 (Graham et al., 1990). At present, one of the most commonly used viability stain combinations is SYBR-14 and PI, sold commercially as LIVE/DEAD Sperm Viability kit (Molecular Probes Inc., OR, USA). When used in combination, the nuclei of living sperm fluoresce green (SYBR-14) and cells that have lost their membrane integrity stain red (PI). This staining technique has been used in a number of species, including the boar (Garner and Johnson, 1995; Saravia et al.,2005, 2007,2009). Although species differences do exist in the function of spermatozoa, the Live/Dead stain may similarly have no adverse affect on fertilization in the equine, although it remains to be tested in this species. Recently a new instrument (Nucelocounter-SP100) has been used to evaluate boar sperm concentration [11]. Due to its compact size and its relatively inexpensive purchase price, this instrument could be useful for field measurements of both concentration and viability. This instrument was considered to be a useful instrument for rapidly measuring stallion sperm concentration and viability (Morrell et al., 2010). Fluorescent probes such as H33258, requiring flow cytometric analysis with a laser that operates in the ultraviolet light range, are less commonly used as this is not a standard feature on the smaller analytical machines. However, one alternative is to use a fluorometer. A fluorometer is a relatively low-cost piece of portable equipment that permits a rapid analysis to be carried out on a sample. Januskauskas et al. (2001) used H33258 to detect nonviable bull spermatozoa by fluorometry and found a negative correlation between the percentage of damaged cells and field fertility. Another option is fluorescent attachments for computer-assisted semen analysis devices. For example, the IDENT fluorescence feature of the Hamilton-Thorne IVOS permits staining with H33258 allowing an assessment of sperm viability to be made along with motility. Fluorochromes used to assess sperm viability by either approach can be used in combination with each other. For example, when CFDA is used along with PI, three populations of cells can be identified: live, which are green; dead, which are red; and a third population which is stained with both and represents dying spermatozoa. Almlid and Johnson (1988) found this combination useful for monitoring membrane damage in frozen-thawed boar spermatozoa during evaluation of various freezing protocols. Harrison and Vickers (1990) also used this combination with a fluorescent microscope and found it to be an effective indicator of the viability of fresh, incubated or cold-shocked boar and ram spermatozoa. Garner et al. (1986) used this combination to stain spermatozoa from a number of species, but at that time could not find a relationship between bull sperm viability detected by CFDA/PI and fertility. Flow cytometry for assessment of sperm viability appears to be a valuable tool for the AI industry. When a high number of sperm is packed in each insemination dose, the effect of selecting the best ejaculates according to sperm viability has a relatively limited effect on NRR56. However, sperm viability might be more important when combined with low-dose inseminations. The FACSCount AF flow cytometer also determines sperm concentration accurately and precisely during the same analysis (Christensen et al., 2004a). The combination of assessment of sperm viability and concentration appears to be useful in the improvement of quality control at AI stations. Because of the results of this trial, this method has been implemented by Danish AI stations (Christensen et al., 2005). Relatively bright fluorescence was found also in the mitochondrial sheath of living sperm. The mechanism by which SYBR-14 binds to the DNA is not known. It is know that PI stains nucleic acids by intercalating betwee n the base pairs (Krishan, 1975). Viability stains have also been used in association with fluorescently labeled plant lectins to simultaneously assess the plasma membrane integrity and the acrosome integrity (Nagy et al., 2003). Assessment of viability using SYBR-14 dye does not damage spermatozoa, since Garner et al. (5) demonstrated that insemination of boar spermatozoa stained with SYBR-14 into sows did not compromise fertilization or the development of flushed porcine embryos in culture. Non-viable cells can be determined using membrane-impermeable nucleic acid stains which positively identify dead spermatozoa by penetrating cells with damaged membranes. An intact plasma membrane will prevent these products from entering the spermatozoa and staining the nucleus. Commonly used examples include phenanthridines, for example propidium iodide (PI; (Matyus, 1984) ethidium homodimer-1 (EthD-1; (Althouse et al., 1995), the cyanine Yo-Pro (Kavak, 2003) and the bizbenzimidazole Hoechst 33258 (Gundersen and Shapiro, 1984). Wilhelm et al. (1996) compared the fertility of cryopreserved stallion spermatozoa with a number of laboratory assessments of semen quality and found that viability, as assessed by flow cytometry using PI, was the single laboratory assay that correlated with stallion fertility. Changes in sperm membrane permeability Detection of increased membrane permeability is employed in different cell types to distinguish different status of membrane organization (Cohen, 1993; Ormerod et al., 1993; Castaneda and Kinne, 2000; Reber et al., 2002). Sperm plasma membrane status is of utmost importance due to its role, not only as a cell boundary, but also for its need for cell-to-cell interactions, e.g. between spermatozoa and the epithelium of the female genital tract and between the spermatozoon and the oocyte and its vestments (for review, see Rodriguez-Martinez, 2001). Membrane integrity and the stability of its semipermeable features are prerequisites for the viability of the spermatozoon (Rodriguez-Martinez, 2006). However, cryopreservation, whose purpose is to warrant sperm survival, causes irreversible damage to the plasma membrane leading to cell death in a large number of spermatozoa (Holt, 2000) or, in the surviving spermatozoa, to changes similar to those seen during sperm capacitation, thus shorten ing their lifetime (Perez et al., 1996; Cormier et al., 1997; Maxwell and Johnson, 1997; Green and Watson, 2000; Schembri et al., 2000; Watson, 2000). During the freezing process, cells shrink again when cooling rates are slow enough to prevent intracellular ice formation as growing extracellular ice concentrates the solutes in the diminishing volume of non-frozen water, causing intracellular water exosmosis. Though warming and thawing, the cells return to their normal volume. Thus, it is important to know the permeability coefficient of the cells to cryoprotectants, as well as the effect of cryoprotective agents on the membrane hydraulic conductivity. Classical combination of probes allows discrimination of two or three subpopulations of spermatozoa, i.e. live, dead and damaged depending on the degree of membrane integrity (Eriksson RodrÄ ±Ã‚ ´guez-MartÄ ±Ã‚ ´nez, 2000). A new, simple and repeatable method to detect membrane changes in all spermatozoa present in a boar semen sample, by use of markers (combination of SNARF-1, YO-PRO-1 and ethidium homodimer) used to track changes in sperm membrane permeability, has been developed recently by our group (Pena et al., 2005). In determined physiological or pathological situations, live cells are unable to exclude YO-PRO-1, but are still not permeable to other dead-cell discriminatory dyes, like propidium iodide or ethidium homodimer. YO-PRO-1 is an impermeable membrane probe and can leak in, only after destabilization of the membrane, under conditions where ethidium homodimer does not. Because several ATP-dependent channels have been detected in spermatozoa (Acevedo et al. , Sperm Assessment Using Flow Cytometry Sperm Assessment Using Flow Cytometry Abstract Flow cytometry is emerging as an important tool in the field of modern andrology for routine analysis of spermatozoa. Recently, application of flow cytometry in the artificial insemination industry especially for pig is a new approach. Until very recent, semen sample analysis was routinely performed by microscopical evaluation and manual techniques by laboratory operators; the analysis is affected by a wide imprecision related to variability among observers, influencing its clinical validity. The last decade, several new flow cytometric techniques have been introduced for farm animal semen assessment that enable a more detailed evaluation of several sperm characteristics. Here in this paper, an initiative has been taken to focus on a number of recent flow cytometry developments important for addressing questions in andrological tests. After the invention of flow cytometry, sperm evaluation by traditional microscopic means became questioned due to the robust advantages of flow cytometry over the microscopic method. Due to the recent development of large number of fluroscence probes, flow cytometry is now capable of analyzing number of sperm characteristics like viability, capacitation, acrosomal integrity, membrane permeability, membrane integrity, mitochondrial status, DNA integrity, decondensation of DNA and differences between gamets based on sex. The application of flow cytometry to their detection allows increased numbers of spermatozoa to be assessed over a short time-period, provides the possibility of working with small sample sizes, increases the repeatability of assessment, removes the subjectivity of assessment and allows simultaneous assessment of multiple fluorochromes. Flow cytometry is a technique capable of generating significantly novel data and allows the design and execution of experiments that a re not possible with any other technique. Nowadays, semen evaluation using laboratory assays is extremely important to the artificial insemination industry to provide the most desired quality product to customers. Future development of flow cytometric techniques will permit further advances both in our knowledge and in the improvement of assisted reproduction techniques. In this paper, the main semen parameters that can be analyzed with fluorochromes and adapted for use with a flow cytometer will be reviewed and the relationship of these tests to fertility will be discussed. Introduction Semen evaluation is the single most important laboratory test that has helped us to identify clear-cut cases of fertility (Jarow et al., 2002), infertility or even of potential sub-fertility (Rodrà ­guez-Martà ­nez, 2007). Determination of the potential fertility of semen sample and, in the long run, of the male from which it has been collected is the ultimate goal of semen evaluations in clinically healthy sires. Methods are available that can sometimes estimate the potential fertilizing capacity of a semen sample and, in some cases, of the male (reviewed by Dziuk 1996; Rodrà ­guez-Martà ­nez et al. 1997a; Rodrà ­guez-Martà ­nez and Larsson 1998; Saacke et al. 1998; Larsson and Rodrà ­guez-Martà ­nez 2000; Rodrà ­guez- Martà ­nez 2000, 2003; Popwell and Flowers 2004; Graham and Mocà © 2005; Gillan et al. 2005). The methods routinely used for evaluation of the quality of a semen sample involved an evaluation of general appearance (i.e. colour, contamination, etc.), volume, pH, sperm concentration, viability, morphology and motility. Most of these techniques are microscopic analyses that only measure a small number of spermatozoa within a population, are time-consuming, can be subjective and generally measure sperm attributes individually. Recently, limitations of semen evaluation methodology have been brought into sharp focus by controversies raised in the epidemiological literature. It should also be noted that such conventional measurements are prone to extreme inter-ejaculate variation, even when the laboratory methodology has been standardized. In the wake of this information, new opportunities have arisen for the development of methods for the diagnosis of male infertility, many of which have been shown to exhibit a prognostic value that eludes conventional semen profiling. Moreover, ejaculated spermatozoa are nowadays handled for use in assisted reproductive technologies, such as the artificial insemination of chilled, frozen-thawed or sexed se men, and IVF. Such handling implies semen extension, fluorophore loading, ultraviolet and laser illumination, high-speed sorting, cooling and cryopreservation, procedures that impose different degrees of change in sperm function following damage to sperm membranes, organelles or the DNA. Therefore, although several assays have been developed to monitor these sperm parameters, recently it is being claimed that buck of these procedures are incomplete, time consuming and laborious. Flow cytometry in different technical applications offers many advantages for the analysis of sperm quality. Flow cytometry allows the simultaneous measurement of multiple fluorescences and light scatter induced by illumination of single cell or microscopic particles in suspension, as they flow very rapidly through a sensing area. The increasing use over the past decade of flow cytometry in the leading laboratories in human and veterinary andrology has dramatically increased our knowledge of sperm function under physiological and biotechnological conditions. Flow cytometers can acquire data on several subpopulations within a sample in a few minutes, making it ideal for assessment of heterogenous populations in semen sample. Initially developed in the 1960s, flow cytometry made automated separation of cells based on the unique recognition of cellular patterns within a population feasible (Hulett et al., 1969). Using such a separation approach, cellular patterns can be identified by as sessing, in individual cells within a population, protein expression using fluorescently labeled antibodies and other fluorescent probes (Baumgarth and Roederer, 2000; Herzenberg et al., 2006). Flow cytometry was first developed for medical and clinical applications such as haematology and oncology. These areas still account for the vast majority of publications on this technique, but during the past few years it has been used in other areas, such as bioprocess monitoring, pharmacology, toxicology, environmental sciences, bacteriology and virology. Recent advancement of flow cytometry increased its application in the reproductive biology especially for andrology. FCM is increasingly used for basic, clinical, biotechnological, and environmental studies of biochemical relevance. Although flow cytometry may overestimate the population of unlabelled cells (Petrunkina and Harrison, 2009), plethora of research from our group in pig (Pena et al., 2003, 2004, 2005; Spjuth et al., 2007; Fernando et al., 2003; Saravia et al.,2005, 2007,2009; De Ambrogi et al., 2006; ) bull (Bergquist et al., 2007; Nagy et al., 2004; Januskauskas et al., 2003; Bergqvist et al., 2007; Hallap et al., 20 05, 2006;) stallion ( Kavak et al., 2003; Morrell et al., 2008) indicate that newly developed fluorescent stains and techniques of flow cytometry has made possible a more widespread analysis of semen quality at a biochemical, ultrastructural and functional level. Therefore, flow cytometry is the current technical solution for rapid, precisely reproducible assessment of sperm suspensions. In this review we have described potentiality and scope of flow cytometry for the evaluation of semen, and the way in which this technique can be used in clinical applications for andrology based on some of our previous experiences. Definition of flow cytometry The definition of a flow cytometer is ‘an instrument which measures the properties of cells in a flowing stream. In other word, a flow cytometer will be defined as ‘an instrument that can measure physical, as well as multi-colour fluorescence properties of cells flowing in a stream. In other work, cytometry refers to the measurement of physical and/or chemical characteristics of cells or, by extension, of other particles. It is a process in which such measurements are made while the cells or particles pass, preferably in single file, through the measuring apparatus in a fluid stream. The data obtained can be used to understand and monitor biological processes and develop new methods and strategies for cell detection and quantification. Compared to other analytical tools, where a single value for each parameter is obtained for the whole population, flow cytometry provides data for every particle detected. As cells differ in their metabolic or physiological states, flow cytometry allows us not only to detect a particular cell type but also to find different subpopulations according to their structural or physiological parameters. Flow cytometry is a technique for measuring components (cells) and the properties of individual cells in liquid suspension. In essence, suspended cells are brought to a detector, one by one, by means of a flow channel. Fluidic devices under laminar flow define the trajectories and velocities that cells traverse across the detector, and fluorescence, absorbance, and light scattering are among the cell properties that can be detected. Flow sorting allows individual cells to be sorted on the basis of their measured properties, and one to three or more global properties of the cell can be measured. Flow cytometers and cell sorters make use of one or more excitation sources and one or two fluorescent dyes to measure and characterize several thousands of cells per second. Flow cytometry gives objective and accurate results (Bunthof et al., 2001; Shleeva et al., 2002), overcoming the problems with the manual methods described above. Function and types of flow cytometry Fluidics, optics and electronics are the three main systems that make up a flow cytometer. In a few minutes, the flow cytometer can acquire data on all subpopulations within a sample, making it ideal for assessment of heterogenous population, such as spermatozoa. The adaptation of flow cytometry to sperm assessment began when it was used for measuring their DNA content (Evenson et al., 1980) and its application to semen analysis has gradually increased over the last 10-15 years. Flow cytometry is now applied to semen evaluation of traits such as cell viability, acrosomal integrity, mitochondrial function, capacitation status, membrane fluidity and DNA status. New fluorescent stains and techniques are continuously being developed that have potential application to the flow cytometric evaluation of spermatozoa. Flow cytometry permits the observation of physical characteristics, such as cell size, shape and internal complexity, and any component or function of the spermatozoon that can be detected by a fluorochrome or fluorescently labeled compound. The analysis is objective, has a high level of experimental repeatability and has the advantage of being able to work with small sample sizes. Flow cytometry also has the capacity to detect labeling by multiple fluorochromes associated with individual spermatozoa, meaning that more than one sperm attribute can be assessed simultaneously. This feature has an added benefit for semen analysis, as few single sperm parameters show significant correlation with fertility in vivo for semen within the acceptable range of normality (Larsson and Rodriguez-Martinez, 2000) and the more sperm parameters that can be tested, the more accurate the fertility prediction becomes (Amman and Hammerstedt, 1993). There are two main types of flow cytometers-analysers and sorters. Sorters have the ability not only to collect data on cells (analyse cells) but also to sort cells with particular properties (defined by the flow cytometer operator) to extremely high purities. There are also a number of commercial flow cytometers that have been developed for particular analytical requirements. Partec manufacture a Ploidy Analyser and also a Cell Counter Analyser. Optoflow has developed a flow cytometer for the rapid detection, characterization and enumeration of microorganisms. Luminex is developing technology for multiplexed analyte quantitation using a combination of microspheres, flow cytometry and high speed digital processing. Advantages of FC compared to other conventional techniques to explore sperm structure and function During the past 2 decades, there has been an increasing interest in reliable assays for assessing semen quality in the fertility clinic and artificial insemination industries. The use of flow cytometry for sperm analysis is an attempt to address the long-standing problem of the subjective nature of the manual method commonly used for semen analysis. An additional source of laboratory variation is the low number of sperms analyzed with manual techniques. Because of time and cost restraints, most laboratories analyze only 50 to 100 sperm to compute the percentage of each cell population and the viability rate. This small sample from a population of millions probably results in a statistical sampling error (Russel and Curtis, 1993). The conventional methods used are limited to microscopic determination of sperm concentration using a hemocytometer (Jorgensen et al., 1997) and evaluation of sperm motility and morphology (Keel et al., 2002). These methods usually involve a subjective asses sment of a few hundred sperm, and quality assurance is rarely implemented in the laboratories performing such analysis. Flow cytometry is a technique that is superior to conventional light microscopy techniques in terms of objectivity, number of cells measured, speed, and precision (Spano and Evenson, 1993). The technique has been used on human sperm to determine a number of factors, including membrane integrity, mitochondrial function, acrosome status, and multiparameter measurement (Garrido et al., 2002). Flow cytometry permitted us to analyze thousands of cells in few seconds. In our series of studies, we demonstrated the feasibility and reproducibility of an automated method to evaluate sperm cell type, count, and viability in human boar samples. In our hand, the precision of the flow cytometric analysis is satisfactory in diverse species (boar, bull, stallion etc), and the observed CVs were significantly better than those reported for the manual method. While there are many advantages of using the flow cytometer for routine semen analysis, its use is often limited to research by the expense and difficulties of operation associated with the requirement of a skilled operator. In addition, a flow cytometer is quite large and cannot withstand shocks associated with movement, meaning it requires a dedicated position in the laboratory. However, the development of more affordable ‘‘bench-top flow cytometers has recently increased the potential application to semen analysis. If we consider flow cytometric analysis further, we can see that it is gaining wider acceptance as a technique for assessing the acrosome reaction and viability simultaneously. Comparing these assays to the more widely used epifluorescent microscopic techniques, the flow cytometric analysis is able to give a far more simple and objective method of analysis, especially with regard to correlation of fertilization with acrosome reactivity potential (Uhler et al., 1993; Purvis et al., 1990; Carver-Ward et al., 1996). A large number of different techniques to estimate sperm concentration have been reported. In the mid-1990s a series of fixed-depth disposable slides were evaluated as rapid and effective pieces of equipment for the estimate of sperm concentration. Preliminary data from a number of studies suggested that, at least in the 20-mm-depth format, such chambers resulted in a noticeable underestimate of sperm concentration compared to the gold standard (improved Neubauer hemocytometer). Using this information, the World Health Organization stated that ‘‘such chambers, whilst convenient in that they can be used without dilution of the specimen, may lack the accuracy and precision of the haemocytometer technique (World Health Organization, 1999). Further data—for example, from Tomlinson and colleagues—showed that 2 proprietary disposable slides (Microcell, Conception Technologies, San Diego, Calif; Leja, Leja Products, BV Nieuw- Vennep, The Netherlands) gave lower spe rm concentrations compared to the hemocytometer method (Tomlinson et al., 2001). To put this in context, numerous reports document unacceptable discrepancies between different laboratories and even between different individuals, although fewer studies attempt to address these issues. So, what is wrong? Several reports emphasize the need for improvement in overall quality of semen testing within and between laboratories (Neuwinger et al., 1990; Jorgensen et al., 1997; Keel et al., 2000). However, the subjective nature of conventional semen analyses, combined with their relatively low precision due to the low number of cells assessed, leads to poor intra- and interlaboratory reproducibility; therefore, the introduction of standardized or quality controlled procedures will probably have a limited effect. The conventional analyses are used to determine whether parameters obtained from an ejaculate are within the range characterized by fertile men, and these methods can therefore provide only unclear cut-off values when used for the prediction of fertility status. Many of the advantages that accrue when using flow cytometry may, when applied to assessment of sperm cells, help overcome some of the mentioned problems found in conventional semen analysis. In the field of semen analysis, validation of a method is important because it is essential to have specific, precise, objective, and accurate laboratory tests to establish a correlation of the data with fertility or to determine the fertility potential of a semen sample correctly (Amann, 1989). Precision of a laboratory test is of great concern to the andrologist in the fertility clinic, since the results of the semen analysis are often used to advise a patient about his fertility and the prognosis for the treatment of the couple. To use established cut-off values and ensure uniform diagnosis, within and between laboratory variations should be determined and followed closely. Accurate determination of sperm cell concentration is critical to the AI industry because it provides assurance both to bull studs and to customers that straws of extended semen contain the sperm numbers indicated. An accurate measure of sperm concentration is particularly important in export markets in which verification of numbers may be required. Routine sperm counts can help to identify possible processing errors within a specific batch of semen or on a particular day, should those errors occur. As sperm counting procedures become more refined, routine counting can be used to monitor subtle changes in daily semen processing that might affect the number of sperm packaged in a straw. Hemacytometers are widely used for routine sperm counts, but the equipment is slow, and multiple measurements of each sample are needed. Single hemacytometer counts are not highly accurate; because of inherent errors in the technique, Freund and Carol (13) found that mean differences of 20% were not uncommon between duplicate sperm count determinations by the same technician. Electronic counters provide much more rapid counting, are easier to use, and give more repeatable results among technicians. However, those instruments tend to include in the sperm count any somatic cells present, immature sperm forms, cytoplasmic droplets, debris, and bacteria, thereby inflating the concentration value (19). Currently, the primary method used by the AI industry to estimate sperm concentration is spectrophotometric determination of turbidity of a semen sample using an instrument previously calibrated for sperm concentration with a hemacytometer or Coulter counter (1). This approach is only as accurate as the methods used for spectrophotometer calibration. New, more accurate methods for sperm count determinations are being sought to replace the older ones. Some laboratories are trying the Maklerm counting chamber (Seif- Medical, Haifa, Israel) and other improved hemacytometers, such as the MicroCellTM (Fertility Technologies, Inc., Natick, MA); however, these techniques will likely have standard lems similar to those associated with the standard hemacytometers. It may be argued that when comparing fluorescent microscopy assays with flow cytometry, one is examining patterns of fluorescence rather than fluorescence intensity, i.e., the flow cytometer is not capable of discriminating sperm which have a fluorescent marker bound to the equatorial segment or over one of the acrosomal membranes (Parinaud et al., 1993; Mortimer and Camenzind, 1989; Mortimer et al., 1987). Tao et al. (1993) compared flow cytometry and epifluorescent microscopy with various lectins and indicated that there is no significant difference between the two methodologies for detection of the acrosome reaction. However, it has been argued that lectins do not bind specifically to the acrosomal region of the sperm (Purvis et al., 1990; Holden and Trounson, 1991) and that other binding sites can be easily distinguished by epifluorescence microscopy, whereas flow cytometry identifies the signal from the entire sperm. Additionally, conventional light microscopic semen assessment is increasingly being replaced by fluorescent staining techniques, computer-assisted sperm analysis (CASA) systems, and flow cytometry (PenËÅ"a et al., 2001; Verstegen et al., 2002). Additional advantages over existing techniques are that this approach is faster than the hemacytometer and that cellular debris, fat droplets, and other particulate material in extended semen are not erroneously counted as sperm, as often occurs with electronic cell counters. This method can also be used to determine the number of somatic cells in a semen sample. Viability The viability of spermatozoa is a key determinant of sperm quality and prerequisite for successful fertilization. Viability of spermatozoa can be assessed by numerous methods, but many are slow and poorly repeatable and subjectively assess only 100 to 200 spermatozoa per ejaculate. Merkies et al. (2000) compared different methods of viability evaluation. They concluded that Eosin-nigrosin overestimate viability while fluorescent microscope and flow cytometry estimate similar trend of viability. Currently flow cytometric procedures have been developed which simultaneously evaluate sperm cell viability, acrosomal integrity and mitochondrial function. This method has been successfully used for assessing spermatozoa viability in men (Garner and Johnson, 1995), bulls (Garner et al., 1994; Thomas et al., 1998), boars (Rodrà ­guez-Martà ­nez, 2007; Garner and Johnson, 1995; Garner et al., 1996), rams (Garner and Johnson, 1995), rabbits (Garner and Johnson, 1995), mice (Garner and Johnson, 1995; Songsasen et al., 1997), poultry and wildfowl (Donoghue et al., 1995; Blanco et al., 2000) and honey bees (Collins and Donoghue, 1999; Collins, 2000) and in fish (Martin Flajshans et al., 2004). Considerable information has accumulated on the use of fluorescent staining protocols for assessing sperm viability (Evenson et al., 1982). The SYBR 14 staining of nucleic acids, especially in the sperm head, was very bright in living sperm. Good agreement was observed between the fluorescent staining method and the standard eosin-nigrosine viability test; the flow cytometric method showed a precision level higher than that of the manual method. One of the first attempts to assess sperm viability utilized rhodamine 123 (R123) to assess mitochondrial membrane potential and ethidium bromide to determine membrane integrity using flow cytometry (Garner et al., 1986). Other combinations that have been used to examine the functional capacity of sperm are carboxyfluorescein diacetate (CFDA) and propidium iodide (PI) (Garner et al., 1988; Watson et al., 1992); carboxydimethylfluorescein diacetate (CMFDA), R123, and PI (Ericsson et al., 1993; Thomas and Garner, 1994); and PI, pisum sativum agglutinin (PSA), and R123 (Graham et al., 1990). At present, one of the most commonly used viability stain combinations is SYBR-14 and PI, sold commercially as LIVE/DEAD Sperm Viability kit (Molecular Probes Inc., OR, USA). When used in combination, the nuclei of living sperm fluoresce green (SYBR-14) and cells that have lost their membrane integrity stain red (PI). This staining technique has been used in a number of species, including the boar (Garner and Johnson, 1995; Saravia et al.,2005, 2007,2009). Although species differences do exist in the function of spermatozoa, the Live/Dead stain may similarly have no adverse affect on fertilization in the equine, although it remains to be tested in this species. Recently a new instrument (Nucelocounter-SP100) has been used to evaluate boar sperm concentration [11]. Due to its compact size and its relatively inexpensive purchase price, this instrument could be useful for field measurements of both concentration and viability. This instrument was considered to be a useful instrument for rapidly measuring stallion sperm concentration and viability (Morrell et al., 2010). Fluorescent probes such as H33258, requiring flow cytometric analysis with a laser that operates in the ultraviolet light range, are less commonly used as this is not a standard feature on the smaller analytical machines. However, one alternative is to use a fluorometer. A fluorometer is a relatively low-cost piece of portable equipment that permits a rapid analysis to be carried out on a sample. Januskauskas et al. (2001) used H33258 to detect nonviable bull spermatozoa by fluorometry and found a negative correlation between the percentage of damaged cells and field fertility. Another option is fluorescent attachments for computer-assisted semen analysis devices. For example, the IDENT fluorescence feature of the Hamilton-Thorne IVOS permits staining with H33258 allowing an assessment of sperm viability to be made along with motility. Fluorochromes used to assess sperm viability by either approach can be used in combination with each other. For example, when CFDA is used along with PI, three populations of cells can be identified: live, which are green; dead, which are red; and a third population which is stained with both and represents dying spermatozoa. Almlid and Johnson (1988) found this combination useful for monitoring membrane damage in frozen-thawed boar spermatozoa during evaluation of various freezing protocols. Harrison and Vickers (1990) also used this combination with a fluorescent microscope and found it to be an effective indicator of the viability of fresh, incubated or cold-shocked boar and ram spermatozoa. Garner et al. (1986) used this combination to stain spermatozoa from a number of species, but at that time could not find a relationship between bull sperm viability detected by CFDA/PI and fertility. Flow cytometry for assessment of sperm viability appears to be a valuable tool for the AI industry. When a high number of sperm is packed in each insemination dose, the effect of selecting the best ejaculates according to sperm viability has a relatively limited effect on NRR56. However, sperm viability might be more important when combined with low-dose inseminations. The FACSCount AF flow cytometer also determines sperm concentration accurately and precisely during the same analysis (Christensen et al., 2004a). The combination of assessment of sperm viability and concentration appears to be useful in the improvement of quality control at AI stations. Because of the results of this trial, this method has been implemented by Danish AI stations (Christensen et al., 2005). Relatively bright fluorescence was found also in the mitochondrial sheath of living sperm. The mechanism by which SYBR-14 binds to the DNA is not known. It is know that PI stains nucleic acids by intercalating betwee n the base pairs (Krishan, 1975). Viability stains have also been used in association with fluorescently labeled plant lectins to simultaneously assess the plasma membrane integrity and the acrosome integrity (Nagy et al., 2003). Assessment of viability using SYBR-14 dye does not damage spermatozoa, since Garner et al. (5) demonstrated that insemination of boar spermatozoa stained with SYBR-14 into sows did not compromise fertilization or the development of flushed porcine embryos in culture. Non-viable cells can be determined using membrane-impermeable nucleic acid stains which positively identify dead spermatozoa by penetrating cells with damaged membranes. An intact plasma membrane will prevent these products from entering the spermatozoa and staining the nucleus. Commonly used examples include phenanthridines, for example propidium iodide (PI; (Matyus, 1984) ethidium homodimer-1 (EthD-1; (Althouse et al., 1995), the cyanine Yo-Pro (Kavak, 2003) and the bizbenzimidazole Hoechst 33258 (Gundersen and Shapiro, 1984). Wilhelm et al. (1996) compared the fertility of cryopreserved stallion spermatozoa with a number of laboratory assessments of semen quality and found that viability, as assessed by flow cytometry using PI, was the single laboratory assay that correlated with stallion fertility. Changes in sperm membrane permeability Detection of increased membrane permeability is employed in different cell types to distinguish different status of membrane organization (Cohen, 1993; Ormerod et al., 1993; Castaneda and Kinne, 2000; Reber et al., 2002). Sperm plasma membrane status is of utmost importance due to its role, not only as a cell boundary, but also for its need for cell-to-cell interactions, e.g. between spermatozoa and the epithelium of the female genital tract and between the spermatozoon and the oocyte and its vestments (for review, see Rodriguez-Martinez, 2001). Membrane integrity and the stability of its semipermeable features are prerequisites for the viability of the spermatozoon (Rodriguez-Martinez, 2006). However, cryopreservation, whose purpose is to warrant sperm survival, causes irreversible damage to the plasma membrane leading to cell death in a large number of spermatozoa (Holt, 2000) or, in the surviving spermatozoa, to changes similar to those seen during sperm capacitation, thus shorten ing their lifetime (Perez et al., 1996; Cormier et al., 1997; Maxwell and Johnson, 1997; Green and Watson, 2000; Schembri et al., 2000; Watson, 2000). During the freezing process, cells shrink again when cooling rates are slow enough to prevent intracellular ice formation as growing extracellular ice concentrates the solutes in the diminishing volume of non-frozen water, causing intracellular water exosmosis. Though warming and thawing, the cells return to their normal volume. Thus, it is important to know the permeability coefficient of the cells to cryoprotectants, as well as the effect of cryoprotective agents on the membrane hydraulic conductivity. Classical combination of probes allows discrimination of two or three subpopulations of spermatozoa, i.e. live, dead and damaged depending on the degree of membrane integrity (Eriksson RodrÄ ±Ã‚ ´guez-MartÄ ±Ã‚ ´nez, 2000). A new, simple and repeatable method to detect membrane changes in all spermatozoa present in a boar semen sample, by use of markers (combination of SNARF-1, YO-PRO-1 and ethidium homodimer) used to track changes in sperm membrane permeability, has been developed recently by our group (Pena et al., 2005). In determined physiological or pathological situations, live cells are unable to exclude YO-PRO-1, but are still not permeable to other dead-cell discriminatory dyes, like propidium iodide or ethidium homodimer. YO-PRO-1 is an impermeable membrane probe and can leak in, only after destabilization of the membrane, under conditions where ethidium homodimer does not. Because several ATP-dependent channels have been detected in spermatozoa (Acevedo et al. , 2006), it seems plausible that this is a result of the silencing of a multidrug transporter. This m