Complement-mediated lysis of pigeon erythrocyte ghosts analysed by flow cytometry. Evidence for the involvement of a 'threshold' phenomenon

Abstract

Flow-cytometric analysis of complement-mediated lysis of antibody-coated pigeon erythrocyte ghosts containing fluorescein was carried out to determine whether lysis involved a gradual release of fluorescein or a 'threshold' release from individual cells. Antibody-coated ghosts were comprised of three subpopulations identified by fluorescence and scatter (size). These were: (a) highly fluorescent, medium scatter, (b) medium fluorescence, high scatter, and (c) low (or zero) fluorescence, low scatter. Lysed ghosts and isolated nuclei were identified by fluorescence microscopy and scanning electron microscopy. Fluorescence distributions analysed by flow cytometry indicated that, after complement attack, those ghosts remaining intact retained all their fluorescent label. A time course of changes in ratios of the three subpopulations indicated that once lysis of an individual ghost was initiated, release of label was complete within 1 min; no stages of intermediary fluorescence appeared, and those ghosts remaining at the end of the experiment retained the same fluorescence intensity as control ghosts. The results supported the hypothesis that complement-mediated cell lysis is a 'threshold' phenomenon; a submaximal response by a cell population representing a complete response by only some of the cells rather than a partial response by all of the cells.