Localization at high resolution of antibody-induced mobilization of vaccinia virus hemagglutinin and the major histocompatibility antigens on the plasma membrane of infected cells

We examined the consequence of simultaneous or independent binding of monospecific antibody to the hemagglutinin (HA) of vaccinia virus and the A-, B- and -determinants of HLA on HeLa or Raji cells or KkDk determinants of H-2 on L929 cells. The bound antibodies were marked by goat-anti-mouse (GAM) or goat-anti-rabbit (GAR) fluorochrome conjugates suitable for light microscopy and GAM or GAR gold conjugates, used in electron microscopy. Specificity and amount of antibody adsorbed was ascertained by complement-mediated lysis of 51Cr-labeled cells and by fluorescence-activated cell sorter analysis. Regardless of the order of either antibody to major histocompatibility complex (MHC) or antibody to HA addition after warming to 37 degrees C, there was evidence by light microscopy for co-patching and co-capping of the viral and host antigens. Electron microscopic examination revealed that goat-anti- rabbit 20 nM gold conjugate and goat-anti-mouse 5 nM gold conjugate, marking respectively the HA and MHC molecules, became concentrated in patched or caps in which the two antigens frequently overlapped or were closely associated. The contiguous MHC and HA antigens were also engulfed, as evidenced from the of two sizes of gold particles inside endocytic vacuoles. The significance of these observations is discussed in relation to the cytotoxic T lymphocyte-mediated killing virus- infected targets.