Page 693 - Clinical Hematology_ Theory _ Procedures ( PDFDrive )

CHAPTER 32 ■ Laboratory Manual: Manual Procedures in Hematology 677

HEMATOLOGY PROCEDURES (continued)

Procedure

1. One thir o a capillary tube shoul be f lle with well-

mixe bloo .

2. An equal amount o f ltere stain is then rawn into the

tube. T e tube is rotate back an orth by han .

3. An alternative metho is to mix two rops o bloo an

two rops o f ltere stain. A

4. Allow this mixture to stan or at least 10 minutes.

5. Gently remix an expel small rops o the stain an

bloo mixture onto several microscope sli es an prepare B

smears.

6. Air- ry.

7. wo or three rie sli es may be counterstaine

with Wright stain (see proce ure or staining bloo

smears). FIGURE 32.5 Miller ocular isc. Square A is nine times the area

8. Using the 10× microscope objective, ocus the smear. A o square B. Reticulocytes are counte in square A; erythrocytes are

a rop o oil to the sli e, an move to the oil immersion counte in square B in successive microscopic f el s until at least

(100×) objective. T e appropriate counting area is the por- 300 RBCs are counte .

tion o the smear where the erythrocytes are evenly is-

tribute an not overlapping. Be ore beginning the count, or anemia is a itionally help ul or clinical interpretation o

scan the sli e to check that reticulocytes can be locate the reticulocyte count (see Chapter 5).

on that sli e.

9. o count the reticulocytes, a minimum o 1,000 (both Miller Ocular Disc

reticulin-containing an nonreticulate ) erythrocytes Principle

must be counte . Normally, 500 erythrocytes will be

counte on each o two sli es. I the number o reticu- A Miller ocular isc inserte into the eyepiece o the micro-

locytes on these two sli es o not agree within 20%, scope permits a rapi survey o erythrocytes. T is isc (see

a thir sli e o 500 erythrocytes must be counte . Be Fig. 32.5) imposes two squares (one nine times the area o the

sure to count all cells that contain a blue-staining f la- other) onto the f el o view.

ment, ragment, or granule o reticulum in the erythro-

cyte. T e counting f el can be re uce by using paper Procedure

hole rein orcers or small pieces o paper cut to f t the Reticulocytes are counte in the large square an erythro-

oculars with a small hole cut out in the mi le o each. cytes in the small square in successive microscopic f el s

his makes counting easier than viewing the entire until at least 300 RBCs are counte . T is allows or an esti-

fel . mate o reticulocytes among a minimum population o 200
erythrocytes. T e absolute reticulocyte count can be eter-
Note: A Miller ocular isc can be use to acilitate counting mine by multiplying the reticulocyte percentage by the

the number o reticulocytes an total RBCs. RBC count.

Calculations Calculations

I 47 reticulocytes are oun when 1,000 erythrocytes are

examine (47 reticulocytes an 953 mature erythrocytes), No o reticulocytes iin large squares
.
×
the reticulocyte count is calculate as ollows: Reticulocytes = 100
×
.
( expresse in percentage ) No o RBCs in small squares 9
47

× 100 4 7.% reticulocytes (uncorrecte )
=
1000 Example: Given that there are 50 reticulocytes in the large
,
squares an 300 re bloo cells in the small squares,

Reporting Results Reticulocytes = 50 100 = 1.886
×
Re erence values: 0.5% to 1.5%; neonates, 2.5% to 6.5%. Some ( expresse in percentage ) 300 ×9

laboratories express the reticulocyte count in absolute rather

than proportional terms. Reporting in absolute terms is Note: Reticulocytes may also be counte by automate

becoming the pre erre metho o reporting. T e correction metho s.

(continued)