Alpha chain crosslinks of human fibrin: purification and radioimmunoassay development for two A alpha chain regions involved in crosslinking.
Rapid and efficient purification methods that include hydrophobic chromatography on Phenyl Sepharose CL-4B have been developed for the peptides that span residues 241-476 (CNBr VIII), and 518-584 (CNBr X) in the A alpha chain of human fibrinogen. Amino acid analysis, NH2-terminal sequence determination, and SDS-PAGE indicated that greater than 95% purity of each peptide was achieved. Sensitive and specific radioimmunoassays capable was achieved. Sensitive and specific radioimmunoassays capable of detecting antigen in the range of 0.1-10.0 pmol/ml have also been developed for CNBr VIII and CNBr X. These assays have been characterized and successfully applied in studies designed to localize the two COOH-terminal A alpha chain regions in column effluents of CNBr-digested fibrinogen and crosslinked fibrin. When these immunoassays were used to study purified preparations of a high molecular weight, crosslink-containing CNBr derivative of a alpha polymer, the data provided immunologic confirmation for the involvement of CNBr VIII and CNBr X in alpha chain crosslinking.
D000595:Amino Acid Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D005337:Fibrin / Q000032:analysis* / Q000378:metabolism; D005340:Fibrinogen / Q000032:analysis*; D006801:Humans; D011863:Radioimmunoassay
D005337:Fibrin; D005340:Fibrinogen