An Improved Direct Method for Obtaining the Total White Cell Count in Avian Blood.

Because of the normal presence of nucleated erythrocytes and thrombocytes in avian blood, it has always been difficult to count accurately all of the white blood cells. For this reason, most direct methods have been based upon some technique providing for the recognition of specific types of cells in the counting chamber and then, from the percentage of these types found in the differential count made upon the stained smear, the total number of white cells per cu. mm. of blood have been calculated. To distinguish between the various types of white blood-cells, previous investigators have used a solution of vital neutral red alone, 1 vital neutral red with formalin in a separate solution, 2 or neutral red with a separate solution containing formalin plus crystal violet. 3 Neutral red, however, stains both granulocytes and monocytes, and it is often difficult to distinguish the one from the other under the powers of magnification that can be used in conjunction with the counting chamber. On the other hand, if one attempts to include both of these cell types in the count, the wide variation in staining intensity of the monocytes frequently occasions inaccuracies through the missing of individual cells. This is especially a source of error when the counting of blood cells is assigned to technicians. Moreover, it was found that the diluting fluids which do not contain a fixative frequently fail to preserve the red blood cells intact, the resulting hemolysis making a further complication in arriving at accurate counts of both the red and white blood cells.

The fact that the predominating granulocyte in avian blood contains eosin-staining granules suggested the use of the dye given below, which, when combined with a fixative to preserve the erythrocytes, stains these granular cells so as to make them stand out sharply and distinctly in the counting chamber.