VLD 411: Veterinary Clinical Biochemistry and Laboratory Diagnosis - I (0+1)

Preparation of Blood smear

PREPARATION OF BLOOD SMEARS
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Blood smears

• It is essential that slides used in making smear preparations be unscratched, non-corroded and meticulously clean, free from grease, dust, acid or alkali; that slides be handled by their edges; that the blood be taken as it exudes; that the process be done rapidly so as to prevent coagulation; and that smears be left to dry in a horizontal position away from flies and dust.
• Mark necessary data with wax pencil on the end of each slide. Blood films should be stained as soon as possible after drying to ensure proper staining.

Preparation of thin blood smear

• A small drop of blood is applied to the slide approximately 20 mm from one end. A spreader (or another clean slide) is placed on the slide at an angle of 20-30o and drawn back to make contact with the blood.
• The blood is allowed to run to each end of the spreader. The blood is spread along the slide in a fairly rapid but smooth motion.
• The spreader should be placed at an angle of less that 45o for making the smear. A thin and even blood smear should result. Wave the slide in the air until it dries (a matter of few seconds if the smear is thin enough).
• If the smear is to be stained in Giemsa’s stain, fix it by dipping in absolute methyl alcohol.
• If the smear is to be stained in Wright or Leishman’s stain, fixation is not necessary since it will take place during the staining process.
• If the smear is to be stored for more than a day or so before staining, it should be fixed in methyl alcohol.
• While Leucocytozoon, microfilariae and sometimes Trypanosoma can be found with the lower powers of the microscope, the stained smears should be examined with the oil immersion objective for other protozoa.
• The faster thin smears dry, the less distortion is produced; hence, the most natural appearing protozoa are at the thin end and around the edges of the smear.

Preparation of thick blood smears

• Use clean slides as for thin smears. Place a medium sized drop of blood or several tiny ones on the slide and mix with a toothpick or the corner of another slide.
• Allow to dry in air or in an incubator at 37^o C; a hair dryer can be used to speed up this process.
• Thick smears must be laked (i.e., the hemoglobin must be extracted) before staining. This can be done by placing them in water until their color has disappeared.
• If Giemsa’s stain is used and the smears are fresh, laking will take place during the staining process. If the smears are to be stored for more than a day or so before staining, they should be laked and then fixed with absolute methyl alcohol before storage, since it is often extremely difficult to remove the hemoglobin from smears that have been stored for some time.