AJP - Cell Physiology, Vol 262, Issue 2 C396-C404, Copyright © 1992 by American Physiological Society
Antibodies against the cystic fibrosis transmembrane regulator
C. M. Fuller, M. B. Howard, D. M. Bedwell, R. A. Frizzell and D. J. Benos
Department of Physiology and Biophysics, University of Alabama, Birmingham 35294-0005.
Rabbit polyclonal antibodies have been raised against high-performance
liquid chromatography purified synthetic peptides corresponding to two
discrete regions of the cystic fibrosis transmembrane regulator (CFTR)
protein: the R-domain (residues 785-796) and the extreme COOH-terminus
(residues 1467-1480). Antibodies (Ab) to each of these peptides were
affinity purified either by passage over a peptide-derivatized agarose
matrix (Ab 785) to produce monospecific polyclonal antibodies or by protein
A affinity chromatography to purify the immunoglobulin G1 fraction free of
other serum proteins (Ab 1467). These antibodies recognize a candidate CFTR
protein in the colonic cell line T84, as determined by Western blot
analysis and by immunoprecipitation and labeling of the precipitate with
[gamma-32P]ATP in the presence of protein kinase A. Both antibodies
precipitated CFTR-related polypeptides from four mammalian cell types
(HeLa, Bsc-40, HEp-2, and Chinese hamster ovary cells) transfected with the
full-length CFTR cDNA clone using a vaccinia T7 protein expression system.
Similar results were observed using a yeast CFTR expression system. In each
case the Mr values of the bands observed were consistent with that expected
for the CFTR protein. These antibodies should be useful probes for the
immunocytochemical localization, immunoaffinity purification, and
ultimately the functional characterization of the CFTR protein.
