Peripheral Smear Using Leishman Stain

PBS using Leishman Stain

1. Peripheral smear

2. Preparations
•Specimen-direct capillary blood or
anticoagulated blood
•Apparatus-clean glass slides,lancet,
spreader slide,microscope&microscope
oil
•Stain-leishman stain,wright or gimsa
•Diluent-distilled water or buffer solution
with Ph 6.8

3. Composition of leishman’s stain •Polychromed methylene blue
•Eosin azure leishman powder
•Acetone free absolute alcohol
•1.5gms leishman powder in 1 liter
acetone free absolute alcohol

4. Procedure
MAKING A SMEAR
•One drop of blood on one end of slide
•Spreader slide is placed at 45°on the drop
and moved along the slide
•It is moved smoothly and once such that
the blood film is thin
•Care should be taken to prevent the
formation of air bubbles
•Air dry the smear
•Make identification mark on one edge

5. Staining a smear
•Place the smear on the staining rack
•Pour leishman stain to cover the smear
completely allow to fix for 2-3 mins
•Add water twice the amount of
leishman’s stain allow to fix for 7-10 mins
•Appearance of golden scum or sheen on
the surface of stain
•Wash the stain off the slide with running
water
•Wipe the back of slide and air dry the
slide

6. Examination of the smear
•The stained slide is placed on the
microscope and is focused under low
power
•Place a drop of oil on the slide and
observe under oil immersion objective
•Observe RBC,platelet series
•WBC series is observed and a differential
count is done
•Record the findings

7. Precautions
•Objective must be clean
•Objective must be cleaned after every use
•Good smear is tongue shaped occupying
2/3
rd
of the slide
•Divided into head,body and tail
•Must be focused at the junction of body
and tail

8. Diagnostic value
RBC’s SERIES
•Size-microcytic/normocytic/macrocytic/
anisocytosis
•Shape-poikilocytosis,sickle cells,target
cells,tear drop cells,burr cells
spherocytosis,ovalocytosis
•Color-hypochromic/normochromic
•Immature cells-
normoblasts,polychromatophillic cells

9. WBC’s SERIES
DIFFERENTIAL COUNT
•Neutrophills-40--70%
•Eosinophills-2--4 %
•Basophills-0--1 %
•Monocytes-2--4 %
•Lymphocytes-25—40 %

10. Immature cells
•Myeloid series-
myeloblasts,promyelocyte,myelocyte,met
amyelocyte,band forms
•Lymphoid series-lymhoblasts

11. Thank you

12. PLATELET SERIES
•General score-6--10platelets/oil
immersion field
PARASITES
•Intracellular-malarial parasite
•Extracellular-microfilarial