High Resolution Immunogold Localization of 'Giardia' Cyst Wall Antigens Using Field Emission SEM with Secondary and Backscatter Electron Imaging

摘要

The report describes the ultrastructural localization of Giardia cyst antigens in the filaments associated with the outer portion of intact cysts and on developing cyst wall filaments in encysting trophozoites. Postembedding immunogold labeling of thin sections of intact Giardia cysts with polyclonal and monoclonal antibodies specific for cyst wall antigens (major protein bands of approximately 29, 75, 88, and 102 kilodaltons on Western blots) showed strong labeling of the filamentous cyst wall, while no labeling was seen on the membranous portion. High resolution field emission scanning electron microscopy (FESEM) of Giardia cysts revealed that the cyst wall specific polyclonal rabbit antisera and monoclonal mouse antibody produced gold labeling of 20 nm filaments in the cyst wall as detected with secondary electron imaging (SEI) and backscatter electron imaging (BEI) at 10 kV, despite coating of the cells with platinum by ion sputtering. FESEM studies of encysting Giardia trophozoites demonstrated that immunostaining with antibodies to cyst wall antigens produced colloidal gold labeling of developing cyst wall filaments on the cell surface; however, the intervening membrane domains were unlabeled. (Copyright (c) 1990 by The Histochemical Society, Inc.)