Production and characterization of monoclonal antibodies to human carbonic anhydrase III.

Abstract

Carbonic anhydrase III (CAIII) is a cytosolic protein found almost exclusively in slow-oxidative skeletal muscle fibers. Upon excessive skeletal muscle activity or damage, CAIII is rapidly released into serum. CAIII is not found in cardiac muscle, whereas the muscle protein myoglobin (Myo) is found in skeletal and cardiac muscle. Because CAIII and Myo are released from injured muscle in a constant ratio, an increase in the Myo/CAIII ratio may be useful as an early diagnostic indicator of acute myocardial damage. Although several reliable Myo immunoassays have been established, no similar CAIII immunoassay is commercially available. We produced murine monoclonal antibodies (MAbs) to human CAIII using standard immunization and cell fusion procedures. Using an enzyme-linked immunoadsorbent assay (ELISA), three MAbs showed strong immunoreactivity with CAIII, but low to moderate levels of cross-reactivity with closely related isoenzymes CAI and CAII. The three MAbs demonstrated unique patterns of reactivity toward CAI, CAII, and CAIII, suggesting that different CAIII epitopes are recognized by the three MAbs. Specificity was further examined by Western blot analysis. These MAbs demonstrated potential for use in the development of an immunoassay for CAIII, and for investigating the biology of skeletal muscle injury in vivo.