Harbor seal (Phoca vitulina) C-reactive protein (C-RP): purification, characterization of specific monoclonal antibodies and development of an immuno-assay to measure serum C-RP concentrations

Vet Immunol Immunopathol. 1997 Oct 6;59(1-2):151-62. doi: 10.1016/s0165-2427(97)00059-7.

Abstract

C-reactive protein (C-RP) was purified from harbor seal (Phoca vitulina) serum by calcium dependant phosphoryl-choline and protein A affinity chromatography. Polyacrylamide gel electrophoresis under reducing conditions revealed a single protein moiety with a molecular weight of approximately 25 kDa. An internal peptide derived from this purified protein was subjected to N-terminal amino acid sequencing. A high amino acid sequence similarity was obtained with other published mammalian C-RP molecules confirming that the purified protein was a C-RP homologue. Eight specific monoclonal antibodies (P13, P51, P87, P101, P106, P130, P157 and P219) were raised against this purified protein. All 8 monoclonal antibodies immunoblotted with the 25 kDa C-RP subunit under reducing conditions. A competitive immunoassay was developed identifying elevated C-RP concentrations in harbor seal serum samples with clinical evidence of inflammatory disease. Application of this immunoassay for the measurement C-RP may provide valuable information for the clinical assessment of harbor seal health.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acute-Phase Reaction / blood
  • Animals
  • Antibodies, Monoclonal / biosynthesis*
  • C-Reactive Protein / analysis
  • C-Reactive Protein / immunology*
  • C-Reactive Protein / isolation & purification*
  • Chromatography, Affinity / veterinary
  • Electrophoresis, Polyacrylamide Gel / veterinary
  • Immunoassay / veterinary*
  • Male
  • Mice
  • Mice, Inbred BALB C
  • Molecular Weight
  • Peptide Fragments / immunology
  • Seals, Earless*
  • Sequence Homology, Amino Acid

Substances

  • Antibodies, Monoclonal
  • Peptide Fragments
  • C-Reactive Protein