Anti-acetylcholine antibodies and first immunocytochemical application in insect brain

A specific immunological approach was developed to enable acetylcholine (ACh) to be visualized in biological tissues. A variety of ACh-like immunogens were synthesized, and injected into rabbits. Antibody specificity was tested using an enzyme-linked immunosorbent assay (ELISA) method. The most immunoreactive ACh derivative was found to be choline-glutaryl-lysine. A mixture of allyl alcohol and formaldehyde was found to be the best fixative of ACh in tissues. The specificity of this antibody recognition was tested in vitro and in immunochemistry. There was excellent agreement between the in vitro results and the ACh staining. Moreover, visualization using these anti-ACh antibodies appeared identical to the results using anti-choline acetyltransferase antibodies.