Hemocyanin-protein A, an immunochemical reagent for scanning and transmission electron microscopy

Abstract

A method is described for preparing hemocyanin-protein A (Hcy/A), a versatile labeling reagent for immunochemical electron microscopy. Because protein A has a specific affinity for the Fc region of immunoglobulins, Hcy/A can be used in indirect labeling with antisera prepared in several different animal species. Because of the distinctive shape and size of hemocyanin, Hcy/A labeling is adaptable to a number of preparative techniques for scanning and transmission electron microscopy. Hcy/A is a stable, highly monomeric conjugate (greater than 90% monomeric, i.e., units of approximately 35 X 50 nm) with a molar ratio of protein A to hemocyanin near unity. Hcy/A labeling is demonstrated using antisera from rabbits, mice and humans to localize cell surface and subcellular antigens. Included are the localization of developmentally expressed erythrocyte antigens, blood group antigens, acetylcholine receptors on Torpedo electroplax membrane fragments, and actin within a subcellular matrix.