FLOW-CYTOMETRY DETECTION OF SURFACE-ANTIGENS ON FRESH, UNFIXED RED-BLOOD-CELLS INFECTED BY PLASMODIUM-FALCIPARUM

The immunofluorescence detection of parasite-specific antigens on the surface of red blood cells infected by Plasmodium falciparum parasites is usually performed by visual detection under a fluorescence microsc ope. We describe here a technique permitting the analysis of surface i mmunofluorescence labelling by flow cytometry. Infected red blood cell s are selected on the basis of their parasitic DNA and RNA content rev ealed by Hoechst and Thiazole Orange vital dyes. Cytometric analysis o f these labels, as well as general erythrocyte characteristics assesse d by analysis of forward and side scatter allows the selection of viab le intact infected erythrocytes from other blood eels. The integrity o f these selected erythrocytes was confirmed by the absence of labellin g with antibodies directed against internal components such as spectri n. This technique permits the detection of specific surface immunofluo rescence staining on red blood cells infected with mature stages of P. falciparum by antibodies in sera from hyperimmune Saimiri monkeys. Us ing Thiazole Orange dye for detection of parasitised cells, this analy sis was performed on a FACSscan apparatus equipped with a single laser .