Hematology Laboratory: Manual RED Cell Counts

CLS 312
 Manual RED Cell Counts

RBC Diluting Fluids

1. Isotonic, so that cells are not hemolyzed or crenated.

2. Must fix the cells to avoid autoagglutination.
3. Hayem's Solution: (HgCl2, NaCl, Na2SO4, H2O)
4. Gower's Solution: (Na2SO4, Acetic Acid, and H2O)
UNOPETTE Procedure : RBC Counts

1. Using the pipet shield of a 10 microliter capillary pipet, open the diaphragm of the dilution vial.

2. Blood from fingerstick or venipuncture is allowed to fill the capillary of the UNOPETTE by capillary action.
Filling of the capillary will stop when it is full.

3. Wipe excess blood carefully from the outside of the

pipet.

4. Using one finger to cover the base of the pipet, insert the pipet tip first into the diluent reservoir while
simultaneously squeezing the container.

5. Release the reservoir and the end of the pipet simultaneously. this will draw the blood sample into the
diluent. Blood is rinsed from the pipet by squeezing and releasing the diluent container several times.

6. Cover the opening at the base of the pipet with a finger and invert the container several times to mix
thoroughly.

7. The base of the pipet can be reinserted into the vial for transportation and may be labelled on the outside.

8. The pipet serves as the dispensing dropper for loading the hemacytometer.

9. Discard the first three drops of mixture from the capillary prior to loading the hemacytometer.

10. Place a clean hemacytometer cover glass over the chamber and allow the chamber to fill by capillary
attraction between the cover glass and the unopette capillary. Count the cells.

11. The dilution ratio for the RBC count is 1:200.

12. Calculations:

# Cells Counted X Dilution Factor X Depth Factor = # cells per cubic millimeter (mm3)
# of Square Millimeters Counted
Interpretation of Results

Normals for a Red Count are as follows:

Women 4.0-5.5 million/mm 3

Men 4.5-6.0 million/mm 3
Newborn 5.0-6.5 million/mm 3

A decreased RBC count is seen in anemia, and an increased count is seen in polycythemia and dehydration.

Sources of Error

Errors in hemocytometry most frequently arise as a result of:

1. apparatus
2. personal technique
3. inherent error

(1) Errors caused by apparatus:

1. chipped pipette tips

2. obscure markings on pipettes
3. non-optically plane cover glasses
4. dirty glassware
5. inaccurate rulings on chamber

(2) Errors caused by personal technique:

1. not thoroughly mixing blood

2. inadequate shaking
3. failure to discard first 4 drops
4. not loading chamber properly (overfilling, trapped air bubbles)
5. counting cells inaccurately (skipping cells, counting cells twice, counting on wrong borders)
6. calculation error
7. clerical error

(3) Inherent errors in hemocytometry include:

1. "field errors" - relates to the random distribution of cells on the counting chamber
2. statistical error - occurs when total number of cells is too low to give statistical confidence in result
(this error is reduced when larger numbers of cells are counted)

47104042.doc
Wednesday, December 01, 2010