(1) New Methylene Blue Solution. Dissolve 0.5 grams of new methylene
blue, 1.4 grams of potassium oxalate, and 0.8 grams of sodium chloride in distilled
water. Dilute to 100 ml. Filter before use.
(2) Brilliant Cresyl Blue Solution. Dissolve 1.0 grams of brilliant cresyl blue
in 99 ml of .85 per cent sodium chloride. Filter before use.
(1) Place 3 or 4 drops of new methylene blue and 3 or 4 drops of blood
(venous or capillary) in a small test tube.
(2) Mix the tube contents and allow to stand for a minimum of 15 minutes.
This allows the reticulocytes adequate time to take up the stain.
At the end of 15 minutes, mix the contents of the tube well.
Place a small drop of the mixture on a clean glass slide and prepare a
Counterstain with Wright's stain, if desired.
Allow smear to air-dry.
(7) Place the slide on the microscope stage and, using the low power
objective, locate the thin portion of the smear in which the red cells are evenly
distributed and are not touching each other.
(8) Switch to oil immersion magnification and count the number of
reticulocytes in 5 fields of 200 RBCs.
Number of reticulocytes counted
= % reticulocytes
e. Sources of Error.
(1) Equal volumes of blood and stain give optimum staining conditions. An
excess of blood causes the reticulum to understain. An excess of stain usually
obscures the reticulum.