A protocol for rapid screening of proteoglycan-degrading metalloproteinase inhibitors.
A procedure is described for rapid, high-volume screening of proteoglycan-degrading metalloproteinase inhibitors. The procedure was developed by modifying the proteoglycan-polyacrylamide particle assay for proteases, polysaccharidases (Nagase and Woessner, 1980), and the 1,9-dimethylmethylene blue (DMB) assay for sulfated glycosaminoglycans (Farndale et al., 1982). The following modifications have made the new protocol suitable for evaluating a large number of compounds simultaneously: 1) Pipetting steps were automated using a pipetting robot. 2) Speed and sensitivity of the 1,9-dimethylmethylene assay were increased by scaling down the assay and measuring absorbance using an automatic multiwell plate reader. 3) Data capture and analysis were simplified using computer programs. The modified procedure is rapid, sensitive, reliable, and requires a small sample size. Above all, the procedure is suitable for high-volume screening: approximately 100 compounds can be evaluated in 1 day.
Baragi
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VM
Parke-Davis Pharmaceutical Research Division, Warner-Lambert Company, Ann Arbor, Michigan 48105.
Jordan
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H
Renkiewicz
R R
RR
D000180:Acrylic Resins; D000818:Animals; D002478:Cells, Cultured; D003627:Data Interpretation, Statistical; D004791:Enzyme Inhibitors / Q000032:analysis; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D008751:Methylene Blue / Q000031:analogs & derivatives; D011509:Proteoglycans / Q000378:metabolism*; D011817:Rabbits; D013997:Time Factors; D000099166:Polyacrylamides
D000180:Acrylic Resins; D004791:Enzyme Inhibitors; D008666:Metalloendopeptidases; D008751:Methylene Blue; D011509:Proteoglycans; D000099166:Polyacrylamides; C016401:1,9-dimethylmethylene blue