Peripheral blood smear preparation

Principle

• A thin wedge (push) film spreads blood so that one area contains a monolayer of cells for morphology and differential counting.

Specimen

• EDTA blood, films made ideally within 1–3 h of collection, or fresh capillary blood without anticoagulant.

Reagents and equipment

• Clean grease-free glass slides, a spreader slide with smooth edge (narrower than the slide), methanol for fixation.

Procedure

• Place a small drop (about 2–3 mm) of well-mixed blood near one end of the slide.
• Hold the spreader at about 30–45° in front of the drop, draw it back to touch the drop and let blood spread along its edge.
• Push forward in one smooth, steady movement.
• Air dry quickly (waving), label with patient details, fix in absolute methanol within about an hour, then stain.

Results

• A good film is about two-thirds to three-quarters the slide length, tapers to a smooth feathered edge, has straight sides not touching slide edges, and no holes or streaks.
• The examination area is just behind the feathered edge where red cells barely touch.

Quality control and pitfalls

• Too thick: large drop, steep angle or fast push; too thin: small drop, low angle or slow push.
• High Hct needs a lower angle; anaemic blood needs a steeper angle.
• Delay in EDTA causes crenation, vacuolation, nuclear changes in WBCs and smudge cells.
• Slow drying causes water artefact (refractile inclusions) and shrunken cells.
• Dirty slides cause holes; uneven pressure causes ridges.
• Large cells (monocytes, blasts) accumulate at edges and tail, so avoid those areas for counting.

Clinical use

• Morphology of RBCs, WBCs and platelets; manual differential; platelet estimate; parasite detection; review of analyser flags.