Double labelling of cell surface antigens with colloidal gold markers

Particles of colloidal gold of different diameters (15 nm and 40) have been used to distinctively label different surface antigens expressed on the surface of human peripheral blood B and T lymphocytes. Silver enhancement has been used to facilitate the observation of the gold particles. Observations were carried out in the backscattered electron imaging mode of the scanning electron microscope. Two different methods have been compared: in Method I the two antigens have been identified by monoclonal antibodies of different classes (IgG and IgM); in Method II monoclonal antibodies of the same subclass were used but the ligands were different (goat anti-murine Ig versus biotin/streptavidin). Some cross-reactivity was observed with Method I, but not with Method II.