Monoclonal Antibodies to and Immunoaffinity Purification of Choline Acetyltransferase from Bovine Brain

Abstract

Three hybridomas producing monoclonal antibodies to bovine brain choline acetyltransferase (ChAT) have been established by fusion of the spleen cells from a mouse immunized with purified enzyme with myeloma NS-1 cells. All three clones produced IgGl antibodies that reacted with enzyme protein denatured with sodium dodecyl sulfate. By using one of the monoclonal antibodies, a rapid and efficient immunoaffinity purification procedure of bovine ChAT has been established. Immunoblot analysis and immunoaffinity purification indicated that bovine ChAT is a single 68-kilodalton protein. The monoclonal antibodies will offer us a good tool to isolate the cDNA clones encoding ChAT.

Antibodies Raised Against Synthetic Peptides React with Choline Acetyltransferase in Various Immunoassays and in Immunohistochemistry

“…By use of the hybridoma technology several groups succeeded in generating monoclonal antibodies against partially purified ChAT from various species (Wainer et al, 1984). A few authors subsequently used these monoclonal antibodies in immunoaffinity chromatography to obtain highly purified enzyme Johnson and Epstein, 1986;Braun et al, 1987;Ishida et al, 1987;Ostermann et al, 1990). In return, such preparations were then successfully used for the production of polyclonal ChAT antibodies.…”

Characterization of Mono‐ and Polyclonal Antibodies Against Highly Purified Choline Acetyltransferase: A Monoclonal Antibody Shows Reactivity in Human Brain

“…Several reports have now been published on ChAT purification using immunoaffinity chromatography with monoclonal antibodies produced after immunization with partially purified enzyme Braun et al, 1987). These preparations have occasionally been used for the production of antisera Johnson and Epstein, 1986;Ishida et al, 1987). Recently we reported on large-scale purification of active ChAT (0.5-I mg/ 10 kg of porcine brain) using immunoaffinity chromatography with a monoclonal antibody selective for porcine brain ChAT (Ostermann et al, 1990).…”