Practical hematology Assist.Prof. Dr.Sajeda Al-Chalabi 1

RBC COUNT HEMOCYTOMETER Hemo: blood Cyto: cell Meter: measurement/counter Thus, it is an instrument used to count the blood cells. 2

It includes: a) Neubauer’s slide b) Cover slip c) RBC pipette d) WBC pipette 3

NEUBAUER’S SLIDE It is the name given to a thick glass slide. In the centre of the slide, there is an H- shaped groove. On the two sides of the central horizontal bar, there are scales for counting the blood cells. The depth of the scales is 1/10mm or 0.1mm. 4

Neubauer Hemocytometer 5

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Hemocytometer Chamber 8

The central square is subdivided into twenty five smaller squares and each of these smaller squares is further subdivided into sixteen smallest squares. These are meant for platelet and RBC counting. 9

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the RBCs are counted in five small squares, four of corners and one of center. (total of 80 smallest squares) 11

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Each of the smaller squares is further subdivided into sixteen smallest squares. Length of one smallest square = 1/5 x 1/4 = 1/20mm Width of one smallest square = 1/20mm Depth of one smallest square = 1/10mm Volume of one smallest square = 1/20 x 1/20 x 1/10 = 1/4000mm³ 13

Cell Counts by Hemocytometer 14

C OUNTING R ULE Do not count cells touching Bottom line Right line Count cells touching the upper& left lines  This is to avoid double counting. 15

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DILUTION FACTORS For RBC counting Blood is filled till mark 0.5 and Hayem’s fluid is then filled till mark 101.Both are thoroughly mixed and then few drops are discarded which contain just the diluting fluid in the stem. Thus, 1 portion out of 101 is discarded. So, 0.5 part of blood is in 100 parts of fluid or, 1 part of blood is mixed in 200 parts of fluid Thus, dilution factor for RBC counting is

C ALCULATION Number of cells/mm³= counted cells in 80 small squares × diluting factor × volume correction factor. =N × 200 ×50=N ×

H AYEM ' S SOLUTION Hayem's solution contains: sodium chloride 0.5%, sodium sulphate 2.5% and mercuric chloride 0.25%. The sodium chloride to prevent hemolyses, the sodium sulphate discourages clumping of the erythrocytes and the mercuric chloride is a preservation 20

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FOCUSING 4X to see the general formation of slide. 10X for WBC counting 40X for RBC counting 22

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THANKS 24