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BMC Pulm Med. 2010 Aug 2;10:40. doi: 10.1186/1471-2466-10-40.

Optimization of total protein and activity assays for the detection of MMP-12 in induced human sputum.

Author information

  • 1Department of Biological Technologies, Pfizer Global Research and Development, 35 Cambridge Park Drive, Cambridge, MA 02140, USA. plapan@wyeth.com

Abstract

BACKGROUND:

Proteolysis of matrix components, in particular elastin, is a major contributing factor to the development of lung diseases such as emphysema and chronic obstructive pulmonary disease (COPD). MMP-12 (macrophage elastase) is a protease known to be involved in the progression of lung disease. The relatively low abundance of MMP-12 has precluded the development of quantitative assays that can accurately measure MMP-12 protein levels and activity across cohorts of healthy and diseased individuals.

METHODS:

Commercial antibodies were screened for performance in sandwich ELISA and capture FRET activity assay formats. Precision, accuracy, sensitivity, dilution linearity, and spike recovery were evaluated using sputum samples.

RESULTS:

Total protein and capture FRET activity assays were developed that were sensitive enough to detect MMP-12 in 37 of 38 donor sputum samples. A comparison of results between the two assays shows that a majority of sputum MMP-12 is in the active form. No differences were seen between normal, asthmatic, and COPD donors.

CONCLUSION:

Sensitive and quantitative assays for both MMP-12 activity and total protein in human induced sputum have been developed. These assays can be used to evaluate MMP-12 as a biomarker for lung disease, and to monitor efficacy of potential therapeutic compounds.

PMID:
20678199
[PubMed - indexed for MEDLINE]
PMCID:
PMC2921351
Free PMC Article
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