Premium
Immunological detection of ECTO‐atpase in chicken and rat tissues: Characterization, distribution, and a cautionary note
Author(s) -
Smith Thomas M.,
Carl Stephanie Ann Lewis,
Kirley Terence L.
Publication year - 1998
Publication title -
iubmb life
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.132
H-Index - 113
eISSN - 1521-6551
pISSN - 1521-6543
DOI - 10.1002/iub.7510450523
Subject(s) - atpase , distribution (mathematics) , microbiology and biotechnology , biology , chemistry , immunology , biochemistry , mathematics , enzyme , mathematical analysis
Abstract We have generated a polyclonal antibody (CKG2) against native chicken gizzard ecto‐ATPase for immunolocalization and immunoprecipitation. Active ecto‐ATPase is immunoprecipitated from solubilized chicken and rat membranes and shown to be localized to the plasma membrane of the chicken smooth muscle cells. This antibody is specific for the ecto‐ATPases, since the more abundant chicken stomach ecto‐apyrase is not recognized in immunoprecipitation, western blot or immunolocalization analyses. The CKG2 antibody cross‐reacts with mammalian (rat) ecto‐ATPase in western blots, with testis being the most abundant source. Interestingly, when the same rat membranes are analyzed by western blot under non‐reducing conditions, the 66 kDa ecto‐ATPase is not recognized, instead a 200 kDa protein is detected, previously postulated to be an oligomer of ecto‐ATPase. However, this 200 kDa cross‐reacting protein is not related to the ecto‐ATPases, but is instead an immunoglobulin binding protein, comprised of 50 kDa subunits.