Beruflich Dokumente
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CLS 312/322
At the completion of this laboratory exercise, the student will be able to:
Upon completion of the lecture on smear examination the student will be able to:
1. Estimate the total WBC count based upon the number of WBC's seen per low power field.
2. Evaluate the distribution of cells seen on the smear to determine if a representative differential may
be performed.
4. Identify the following cell types and state their function and normal peripheral blood values:
a. Neutrophil
b. Basophil
c. Eosinophil
d. Polychromatophilic erythrocyte
e. Platelet
f. Monocyte
g. Lymphocyte
6. List and explain how to prevent 3 conditions which would lead to artifactual changes in red cells
resulting in crenation.
7. List and explain how to prevent 3 conditions which would lead to artifactual changes in red cells
resulting in schistocytes.
NORMAL DIFFERENTIALS
Procedure
- In the feathered edge, where the RBC's lay side by side (just touching one another), count 100
white blood cells.
- Use the oil immersion lens estimate the number of platelets per field
- Look at 5-6 fields and take an average
- Multiply the average by 20,000
- Note any macroplatelets
- smudge cells
- eosinophil with no cytoplasmic membrane and with scattered granules
- Pyknotic cell (nucleus extremely condensed and degenerated, lobes condensed
into small, round clumps with no filaments interconnecting).
- Basket cells
b. If a cell does not fit into a category or you are unsure of what the cells is, ask someone to look at it.
(may be abnormal or immature)
c. Make sure to count in an area where cells are distributed evenly and the central pallor of RBC's is
present. RBC's should touch or only slightly overlap. The pattern of scan in the diagram below is
not to scale. It is intended to show direction of movement only. (ref: NCCLS Standards, Fig.1 p. 14
Sec.5.6.2 Vol. 12 No. 1)
d. Always check the edge of a smear for platelets if the platelet estimate is decreased.
g. Always check WBC morphology. Don't assume the diff is finished after classifying 100 cells.
Depending on your findings, you might have to count 200 cells. See section on Abnormal
Differentials.
a. MCV = Hct x 10
RBC
c. Normals:
Females: 88 +/- 9 fl
Males: 87 +/- 7 fl
Microcytic,
hypochromic
normochromic
hyperchromic
Normocytic,
hypochromic
normochromic
hyperchromic
Macrocytic,
hypochromic
normochromic
hyperchromic
e. N,N = 33 - 36% saturated with Hgb. This is the maximum level; if higher would precipitate
out, therefore RBC's cannot actually be hyperchromic.
SIZE
HEMOGLOBIN CONTENT :
SHAPE
ARTIFACTS
If spiculated RBC's are a true feature, they are usually uniformly distributed and do not affect most of the
red cells. However, it is advisable to repeat on a fresh sample.
TARGET CELLS
Very easy artifact to make (dehydration environment) May be caused by contamination with dirt or
alcohol on slide, finger, needle or tube. Usually artifactual target cells will be unevenly distributed
---present in some areas of the smear and absent in others. True target cells are usually evenly distributed
throughout the smear.
DRYING (MOISTURE) ARTIFACTS:
Pseudo-Hypochromia Pale central area has a sharp demarcation rather than gradual transition in color
intensity. Sometimes drying artifact appears as vacuoles or small bubbles which are refractile areas in
the RBC's that will appear to change in size (grow larger or smaller) as the focus in changed-this type is
probably more stain-related than preparation related. Crenated (Moth-eaten) periphery sometimes present.
CAUSED BY:
Humidity
Moist slides
Anemia (increased amounts of plasma to red blood cells)
Pernicious anemia and folate deficiency with
low hematocrits frequently demonstrate this.)
Under-fixation.
Water in alcohol fixation (hygroscopic absolute methyl
alcohol in bottle absorbs water vapor if not
kept tightly closed).
Excess buffer to stain.
Thick smears (and thick area of smear)
ELIMINATE BY:
Avoid Humidity
Air conditioning
Smears made in humid-free lab
Desiccator or Dehydrating Set-up
Calcium Chloride crystals
Smear in desiccator for one hour
Examine red cells in thin area of smear
Is MCHC Compatible?
FRAGMENTS
Thin area-Spherocytes which are really "spheroidocytes" or flattened red cells. True spherocytes will
be found in other (Good) areas of smear.
A normal red blood cell should be approximately the same size as a normal lymphocyte nucleus or 2
normal sized red blood cells should fit side by side across a normal sized poly (not a hypersegmented
poly).
1+ 2+ 3+ 4+
1-6 per oil imm. field 7-10 per OIF 11-20 per OIF > 20 per OIF
Where possible use macrocytic and microcytic, rather than simply anisocytosis alone, when describing
red cell morphology.
Use specific cell morphology when possible, rather than simply reporting poikilocytosis.
When red cells are normocytic, normochromic, report out as NORMAL. When abnormal morphology
has been noted, DO NOT indicate normal on the report form.
ABNORMAL DIFFERENTIALS
a. WBC > 15.0 (>20.0 for babies under 1 month and labor unit)
b. Three or more basophils seen.
2. If more than five immature WBC's are seen (or any blasts) let someone else diff slide and average
results.
3. Correct WBC for NRBC's if you seen ten or more nrbc's/100 WBC.
5. If any cell type is extremely elevated (such as bands, monos, or eos > 20) indicate that you are aware
of the abnormality by circling or checking on the card next to the results.
Examination of the Peripheral Blood Smear: Common Findings Associated with Disease
41900004.doc
Wednesday, October 13, 2010