Manual of basic techniques for a health laboratory - libdoc.who.int

9. Haematology 317Fig. 9.108 Preparing the cresylblue solutionFig. 9.109 Collecting a capillary blood sample9.12.3 Method1. Filter a little of the cresyl blue solution into a test-tube. In the bottom of anothertube place two drops of filtered cresyl blue solution (Fig. 9.108).2. Collect a few drops of blood from the patient’s finger with a Pasteur pipette (Fig.9.109), or use venous blood collected in EDTA dipotassium salt solution andmix well.3. Add two drops of blood to the tube containing cresyl blue solution.4. Mix by gently shaking the tube. Plug the tube with non-absorbent cotton wool.Leave for 15 minutes.5. Take the tube and shake it gently. Remove one drop of the mixture. Place it ona slide ready for spreading.6. Make a thin smear of the mixture with the spreader (see section 9.10.3).Leave the smear to air-dry.9.12.4 Microscopic examinationExamine the smear using the ¥ 100 oil-immersion objective (Fig. 9.110). Look atthe end of the smear, where the erythrocytes should be well separated from eachother. Erythrocytes stain pale blue.Examine at least 100 erythrocytes. Keep a careful count of the total numberof erythrocytes examined and the number of these that are reticulocytes.(Counting is easier if the size of the microscope field is reduced. This can bedone by placing in the eyepiece a small circular piece of stiff black paper in whicha hole of about 5mm in diameter has been punched.)Some haematologists prefer reticulocytes to be reported in terms of the numberconcentration (number of reticulocytes per litre of blood), while others preferthem to be reported in terms of the number fraction (the proportion oferythrocytes that are reticulocytes). Depending on the practice in yourlaboratory or the specification of the requesting physician, make the appropriatecalculation. 11Traditionally, reticulocytes have been reported in the form of percentages (i.e. the proportion,expressed as a percentage, of the erythrocytes that are reticulocytes). If 500 erythrocytes areobserved on the blood film and n of them are reticulocytes, the percentage of erythrocytes iscalculated by multiplying n by 0.2.Example:Of 500 erythrocytes examined, 25 are reticulocytes. The percentage of reticulocytes is then25 ¥ 0.2 = 5%. The normal range for newborn infants is 2.0–6.0%, and that for adults and childrenis 0.2–2.0%.