Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
7632722 | Journal of Pharmaceutical and Biomedical Analysis | 2012 | 7 Pages |
Abstract
This paper summarizes the development and validation of five enzyme activity methods to assess the specific inhibition of human endogenous matrix metalloproteinases MMP-1 (interstitial collagenase), MMP-2 (gelatinase A), MMP-3 (stromelysin 1), MMP-8 (collagenase 2) and MMP-13 (collagenase 3) by anti-Collagenase Clostridium histolyticum (CCH) antibodies in human serum. These MMPs are of interest since antibodies against a therapeutic enzyme may cross-react with, and inactivate, the MMPs. The validated methods utilize spiked exogenous individual MMPs added to serum to determine if the serum inhibits MMP enzyme activity. Factors evaluated and optimized during development include pH, reaction time and temperature, inhibitor concentration for the positive control, and substrate and serum concentration. Characteristics established during validation for each MMP activity inhibition method included intra- and inter-assay precision and recovery, recovery in the pooled normal human serum samples, bench-top stability at room temperature and on wet ice, and assay cut-point determination. Precision results ranged from â¼1 to 12% CV, recoveries of the activities of the exogenous MMPs ranged from â¼84 to 90% and cut-point values ranged from 67 to 91%.
Keywords
Related Topics
Physical Sciences and Engineering
Chemistry
Analytical Chemistry
Authors
Thomas J. Edkins, Jack A. Alhadeff, Vincent Kwok, Charles Kalensky, Marie T. Rock, Thomas J. Vidmar, Benjamin J. Jr.,