TY - JOUR
T1 - Membrane antigens in chronic active hepatitis
T2 - Physicochemical and immunological studies of soluble liver homogenate fractions
AU - Shelton, Linda L.
AU - Lee, William M.
AU - Curtis, G. Michael
AU - Arnaud, Philippe
AU - Galbraith, Robert M.
N1 - Funding Information:
I This work was presented in part at the Annual Meeting of the American Association for the Study of Liver Diseases, Chicago, November 1981, and published in abstract form (Hepatology 1, 586, 1982). This research was supported by a grant from The Houghton Foundation and by State of South Carolina Biomedical Research Grants 22230-A210 and 22230-GR22 and from South Carolina state appropriated funds administered through the College of Graduate Studies, Medical University of South Carolina, where L. Shelton is a graduate student. This is publication 587 from the Department of Basic and Clinical Immunology and Microbiology. * To whom requests for reprints should be addressed: Gastroenterology Division, Medical University of South Carolina, 171 Ashley Ave., Charleston, S.C. 29425.
PY - 1983/7
Y1 - 1983/7
N2 - Since the liver membrane components termed liver-specific protein (LSP) and liver membrane antigen (LM-Ag) have been implicated as target antigens in chronic active hepatitis (CAH), we performed physicochemical and immunological analyses of these and other fractions obtained from liver homogenates. Supernatants were prepared from homogenates of human and rabbit liver by centrifugation at 105,000g for 1 hr; after gel filtration on Sepharose 6B, the LSP fraction was obtained as the excluded peak I and LM-Ag as an included fraction, peak IIB. Comparison by immunodiffusion with reference LSP and guinea pig antisera to LSP confirmed the similarity of our fractions to reference preparations. Upon SDS-polyacrylamide gel electrophoresis (SDS-PAGE), human and rabbit peak I were found to contain at least 25 distinct protein bands with a wide range of molecular weights (14,000->100,000) and the other peaks were found to be equally heterogeneous. Thin-layer polyacrylamide gel isoelectric focusing (PAGIF) revealed 35-40 separate bands in both peaks I and IIB with a wide range of isoelectric points (3.5-10.0). These results indicate that liver homogenate fractions such as LSP and LM-Ag can no longer be considered as single protein preparations, and are in fact highly complex mixtures of multiple proteins. Further separation and characterization of such fractions will facilitate identification of liver antigens for use in in vitro immunological assays.
AB - Since the liver membrane components termed liver-specific protein (LSP) and liver membrane antigen (LM-Ag) have been implicated as target antigens in chronic active hepatitis (CAH), we performed physicochemical and immunological analyses of these and other fractions obtained from liver homogenates. Supernatants were prepared from homogenates of human and rabbit liver by centrifugation at 105,000g for 1 hr; after gel filtration on Sepharose 6B, the LSP fraction was obtained as the excluded peak I and LM-Ag as an included fraction, peak IIB. Comparison by immunodiffusion with reference LSP and guinea pig antisera to LSP confirmed the similarity of our fractions to reference preparations. Upon SDS-polyacrylamide gel electrophoresis (SDS-PAGE), human and rabbit peak I were found to contain at least 25 distinct protein bands with a wide range of molecular weights (14,000->100,000) and the other peaks were found to be equally heterogeneous. Thin-layer polyacrylamide gel isoelectric focusing (PAGIF) revealed 35-40 separate bands in both peaks I and IIB with a wide range of isoelectric points (3.5-10.0). These results indicate that liver homogenate fractions such as LSP and LM-Ag can no longer be considered as single protein preparations, and are in fact highly complex mixtures of multiple proteins. Further separation and characterization of such fractions will facilitate identification of liver antigens for use in in vitro immunological assays.
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U2 - 10.1016/0090-1229(83)90197-6
DO - 10.1016/0090-1229(83)90197-6
M3 - Article
C2 - 6872359
AN - SCOPUS:0020510345
SN - 0090-1229
VL - 28
SP - 135
EP - 141
JO - Clinical Immunology and Immunopathology
JF - Clinical Immunology and Immunopathology
IS - 1
ER -