The C terminus of the anaphylatoxin C3a generated upon complement activation represents a neoantigenic determinant with diagnostic potential.
Activation of the C component C3 results in generation of the anaphylatoxin C3a. The C3a polypeptide chain consists of 77 amino acids. The active site of this potent mediator, which also has immunoregulatory function resides in its C terminus. This report demonstrates that the C terminus of C3a (C3a-desArg) exposed by proteolytic cleavage from C3 represents a neoantigenic determinant. Two mAb specific for this epitope were obtained after immunization with the synthetic octapeptide (OP) Arg-Ala-Ser-His-Leu-Gly-Leu-Ala [C3a(69-76)] coupled to the carrier keyhole limpet hemocyanin (KLH). These anti-C3a(69-76) antibodies (H453 and H454) reacted in an ELISA system with C3a and KLH-OP but not with C3 or with KLH alone. Free OP efficiently blocked binding of the antibodies to C3a, whereas binding of another anti-C3a mAb (H13) remained unaffected. In immunoblotting analysis, the anti-C3a(69-76) mAb reacted with purified C3a but failed to react with the denatured, noncleaved C3. A novel quantitative C3a-ELISA was established with the anti-C3a(69-76) mAb. It had a sensitivity in the nanogram range (1 to 5 ng/ml). The C3a determination was not impaired by the presence of high concentrations of C3. Therefore, C3 removal was not required in contrast to the previously described C3a assays. This C3a ELISA might facilitate clinical C3a quantitation, e.g., in samples from patients with adult respiratory distress syndrome. In these patients, C3a determination in the early phase of the disease is of diagnostic relevance and has prognostic value.
Burger
R
R
Institute for Immunology, University of Heidelberg, Federal Republic of Germany.
Zilow
G
G
Bader
A
A
Friedlein
A
A
Naser
W
W
D000595:Amino Acid Sequence; D000706:Anaphylatoxins / Q000096:biosynthesis / Q000276:immunology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis; D000918:Antibody Specificity; D000937:Antigen-Antibody Reactions; D003167:Complement Activation; D003176:Complement C3 / Q000096:biosynthesis / Q000276:immunology; D015926:Complement C3a; D000939:Epitopes / Q000276:immunology; D007118:Immunoassay; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000138:chemical synthesis / Q000276:immunology; D010452:Peptide Biosynthesis; D010455:Peptides / Q000276:immunology
D000706:Anaphylatoxins; D000911:Antibodies, Monoclonal; D003176:Complement C3; D000939:Epitopes; D009842:Oligopeptides; D010455:Peptides; D015926:Complement C3a