Antisera to gamma-aminobutyric acid. I. Production and characterization using a new model system.

Abstract

Antisera to the amino acid gamma-aminobutyric acid (GABA) have been developed with the aim of immunohistochemical visualization of neurons that use it as a neurotransmitter. GABA bound to bovine serum albumin was the immunogen. The reactivities of the sera to GABA and a variety of structurally related compounds were tested by coupling these compounds to nitrocellulose paper activated with polylysine and glutaraldehyde and incubating the paper with the unlabeled antibody enzyme method, thus simulating immunohistochemistry of tissue sections. The antisera did not react with L-glutamate, L-aspartate, D-aspartate, glycine, taurine, L-glutamine, L-lysine, L-threonine, L-alanine, alpha-aminobutyrate, beta-aminobutyrate, putrescine, or delta-aminolevulinate. There was cross-reaction with gamma-amino-beta-hydroxybutyrate, 1-10%, and the homologues of GABA: beta-alanine, 1-10%, delta-aminovalerate, approximately 10%, and epsilon-amino-caproate, approximately 10%. The antisera reacted slightly with the dipeptide gamma-aminobutyrylleucine, but not carnosine or homocarnosine. Immunostaining of GABA was completely abolished by adsorption of the sera to GABA coupled to polyacrylamide beads by glutaraldehyde. The immunohistochemical model is simple, amino acids and peptides are bound in the same way as in aldehyde-fixed tissue and, in contrast to radioimmunoassay, it uses an immunohistochemical detection system. This method has enabled us to define the high specificity of anti-GABA sera and to use them in some novel ways. The model should prove useful in assessing the specificity of other antisera.

GABAergic neurons in the rat hippocampal formation: ultrastructure and synaptic relationships with catecholaminergic terminals

“…In tests for specificity, the immunoreactivity visualized in sections of tissue was not blocked by 10 mM unconjugated GABA or 100 PM BSA-conjugated glutamate, /3-alanine, or taurine but was completely abolished by preincubation with 10 PM GABA-BSA (Lauder et al, 1986). The antiserum specificity was similar to that described previously by Hodgson et al (1985).…”

Immunogold electron microscopic demonstration of glutamate and GABA in normal and deafferented cerebellar cortex: correlation between transmitter content and synaptic vesicle size.

“…Three primary antisera were used: anti-GABA characterized by Hodgson et al (18), anti-glutamate characterized by Heplcr et al (16), and anti-glycine characterized by Van den Pol and Gorcs (35). Adjacent ultra-thin sections for staining for GABA (dilution li800).…”

Proceedings of the Physiological Society, 14‐15 June 1985, St Andrews Meeting: Communications

“…The specificity of this antiserum has been thoroughly tested and it has been used to selectively visualize GABAergic neurones in the C.N.S. (Hodgson, Penke, Erdei, Chubb & Somogyi, 1985). Thus it seems likely that the neurones detected in the present study are enteric GABAergic neurones.…”