Ji. Moreno et al., IMMUNOCYTOCHEMICAL LOCALIZATION OF THE BETA(2) SUBUNIT OF THE GAMMA-AMINOBUTYRIC ACID(A) RECEPTOR IN THE RAT-BRAIN, Journal of comparative neurology, 350(2), 1994, pp. 260-271
An antiserum to the beta(2) subunit of the rat gamma-aminobutyric acid
(GABA(A)) receptor was prepared by immunizing a rabbit with a fusion
protein expressed in bacteria. The fusion protein had the large, intra
cellular loop expanding between the putative M3 and M4 transmembrane d
omains of the beta(2) subunit fused to staphylococcal protein A (SPA).
The antiserum immunoprecipitated both the solubilized and the affinit
y-purified GABA(A) receptors. The anti-beta(2) antibodies were affinit
y purified on immobilized beta(2) intracellular loop peptide. The anti
bodies recognized a 55-57 kDa peptide in immunoblots of either crude m
embranes from rat cerebral cortex or affinity-purified GABA(A) recepto
rs from bovine cerebral cortex. Immunocytochemistry with the affinity-
purified antibody has revealed for the first time the localization of
the beta(2) subunit in the rat brain. A comparative study of the regio
nal and cellular immunoreactivities of the affinity-purified anti-beta
(2) antibody and the monoclonal antibody 62-3G1 (which recognizes both
beta(2) and beta(3) subunits) is presented. The procedure described f
or generating and preparing specific anti-beta(2) subunit antibodies t
hat are valuable for immunocytochemistry could be extended to other GA
BA(A) receptor subunits. (C) 1994 Wiley-Liss, Inc.