Azure B-eosin staining of blood cells: the effects of variation in stain formulation and staining technique on stain performance

A quantitative study of azure B-eosin-stained blood cells is reported. The effects of variation in stain formulation and staining technique on the binding of azure B and eosin by acidophilic, basophilic and neutrophilic substrates were measured by scanning microdensitometry. The variables considered were stain concentration (the azure B-eosin concentration ratio was constant), azure B concentration (at constant eosin concentration), eosin concentration (at constant azure B concentration), staining time, buffer pH, metal salt contamination, dye contamination, buffer concentration and fixation time. The last two were the only variables which failed to produce changes in dye uptake. Tentative explanations of the observed effects are advanced.