Rapid identification and detection of parasitized human red cells by automated flow cytometry

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Abstract
Rapid and reliable identification of various human red cell parasites is important in many chemotherapeutic and immunologic studies. Because manual microscopic counting is tedious and imprecise, we have developed a simple diagnostic procedure for the automated flow cytometric detection of in vitro infected red cells, using a nucleic acid‐binding fluorescent dye, acridine orange. Human malaria (Plasmodium falciparum)‐infected red cells from continuous human erythrocyte culture were incubated at room temperature in acridine orange stain for 5 min after which the sample were analyzed by flow cytometry. Since mature red cells contain no DNA, infected red cells were identified with a distinct fluorescent signal. A total of 200,000 cells per sample were counted and analyzed in less than 2 min. Rings, trophozites, and schizonts were assessed and identified in synchronozed infected red cell cultures by flow cytometry. In addition, various stages of infected red cells were isolated with a cell sorter. This rapid method permits accurate and reliable assessment of data with the exclusion of anomalous data such as damaged cells, extraneous material, and contaminating particles.