Antigen specificity and cross-species reactivity of a monoclonal antibody (mAb 72.11) against porcine pancreatic procolipase.
We have studied the antigen specificity and cross-reactivity of a monoclonal antibody (mAb 72.11) of subclass IgG1, raised against the precursor form of porcine colipase (procolipase), whose epitope lies near the amino terminal region of the polypeptide. mAb 72.11 cross-reacts with native porcine, equine and human procolipase, as shown by immuno-inactivation and ELISA titration studies carried out on pure proteins, pancreatic tissue homogenate or pancreatic juice. The epitope site recognized by mAb 72.11 was further characterized by studying antibody binding to denatured procolipase. Reduced carboxymethylated procolipase reacted with mAb 72.11 in ELISA. Heat inactivated or reduced carboxymethylated porcine procolipase displaced antigen from the complex formed between antibody and native procolipase. The lack of sensitivity of epitope recognized by mAb 72.11 on procolipase to heat denaturation or reduction of the disulfide bridges is indicative that antigen specificity of mAb 72.11 is not dependent on the conformation of the antigenic site. Cross-reactivity of mAb 72.11 with procolipase from the three species demonstrates that substitution of amino acid at positions 1 and 3 causes no loss of antigenicity. Finally, mAb 72.11 was coupled to sepharose to isolate human procolipase from human pancreatic juice and to separate the precursor form from activated colipase non-adsorbed on the column.
Dezan
C
C
Institut de chimie biologique, faculté des sciences St Charles, Marseille, France.
Rugani
N
N
de la Fournière
L
L
Sarda
L
L
Bellon
B
B
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000302:isolation & purification; D000936:Antigen-Antibody Complex; D002846:Chromatography, Affinity; D003089:Colipases / Q000032:analysis / Q000276:immunology* / Q000378:metabolism; D003429:Cross Reactions; D004151:Dipeptides / Q000494:pharmacology; D004789:Enzyme Activation; D004792:Enzyme Precursors; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes; D006736:Horses; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology; D008969:Molecular Sequence Data; D010179:Pancreas / Q000201:enzymology; D011498:Protein Precursors / Q000032:analysis / Q000276:immunology*; D021382:Protein Sorting Signals; D013552:Swine
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D003089:Colipases; D004151:Dipeptides; D004792:Enzyme Precursors; D000939:Epitopes; D007074:Immunoglobulin G; D011498:Protein Precursors; D021382:Protein Sorting Signals; C071482:calpeptin; C037107:procolipase; C058076:calpain inhibitor III