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STAINING THE CELLS.-Once a blood smear
is made, it should be stained. Staining the blood smear
highlights the differences among the
different types of leukocytes for
easier recognition during the counting
process. The most popular stain used for this purpose
is Wright's stain. Wright's stain is a
methyl alcohol (methanol) solution of
an acid dye and a basic dye. The acid
dye in Wright's stain is known as eosin and is red
in color. The basic dye in Wright's stain is known as
methylene blue and is blue in color.
Generally, white cells are identified
by their affinity to the dye they
prefer. For example, cells that prefer the acid dye
(eosin) are called eosinophils. Other cells
that prefer the basic dye are called basophils.
WARNING Wright's staining solution contains methanol, which is considered a hazardous material. It is classified as flammable, a poison, and an irritant. Methanol must be kept away from heat, sparks, and open flames. Good ventilation in usage areas is paramount since exposure to vapors can irritate eyes, nose, throat, and mucous membranes of the upper respiratory tract. When not in use, methanol containers should be closed tightly and stored upright to prevent leakage. Gloves and protective clothing (e.g., lab coat or apron) and eyewear should be worn to avoid contact with the solution. Absorption through skin can cause permanent blindness. Death may result from ingestion or exposure to high vapor concentrations of methanol. There are a variety of staining products on the market today. Some of these staining products have combined Wright's solution with other staining solutions, such as Giemsa stain. When using a new product, you should always review the manufacturer's usage and safety recommendations. The staining process that we will cover in this chapter is known as a quick stain. A quick stain has very few equipment requirements and only a few procedural steps. An example of a quick stain is One Step II Wright-Giemsa Stain Solution(r) by Criterion Sciences. To stain a blood smear with this product, follow the steps below. 1. Prepare two staining containers by filling one with One Step II stain solution and the other with deionized or distilled water. The use of tap water instead of deionized or distilled water is not recommended since the pH of tap water varies. If tap water is used, its pH should between 5.8 and 7.03. 2. Immerse the slide (blood smear) in the stain for 15 to 30 seconds. (To prevent debris or precipitate from contaminating the slide, do not add new stain to old.) 3. Remove the slide and allow excess stain to drain from the edge of the slide. 4. Immerse the slide in the deionized or distilled water for 5 to 15 seconds. (Change the water when it becomes dark blue or when film forms on the surface.) NOTE: Rinse time is critical and must be shorter than the stain time.
5. Drain excess water and wipe the back of the slide to reduce background color. 6. Place slide in horizontal position on table and allow to air dry. NOTE: Do not accelerate drying time by placing slide on a warmer or in front of a fan. The film of water on the slide is important for the color development. 7. Once the slide is dry, proceed to step 3, counting the cells. |
 
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