The microscopic double immunofluorescence technique, a method for quantitative differentiation between extra- and intracellularly located bacteria in isolated polymorphonuclear granulocytes.

A simple microscopic double immunofluorescence staining technique for the study of phagocytosis of living bacteria by human polymorphonuclear granulocytes (PMN) is presented. This technique enables the differentiation between cell-attached and ingested bacteria and allows the kinetics of phagocytosis to be monitored. The practicability and accuracy of this method is demonstrated by using two genetically defined Yersinia enterocolitica strains of different virulence, and isolated PMN resuspended in serum-free and serum-containing medium, respectively. The advantages of this method over other techniques are discussed.