A semiautomated, 96-well plate assay for collagen synthesis.
We have established a rapid method for the measurement of collagen synthesis in large numbers of cell cultures. 3H-labeled collagen in microwell cultures was salt precipitated, harvested, and washed using a commercially available cell harvester and the filtered collagen was directly counted. The cell number could then be assessed either by methylene blue staining or by dissolving the cells in NaOH and estimating the protein content with bicinchoninic acid. In all of these procedures, the samples remain in microtiter plates, thus ensuring that minimal amounts of reagents are used and minimizing the amount of manipulation necessary. The intergroup variability is 3 to 9% and the recovery of 3H-labeled collagen is greater than 90%. Using this method, large numbers of samples can be assessed for collagen synthesis quickly, conveniently, and for minimal cost.
Scutt
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A
Gesellschaft für Biotechnologische Forschung mbH, Abteilung Genetik, Braunschweig, Germany.
Berg
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A
Mayer
H
H
D000818:Animals; D001769:Blood; D002452:Cell Count; D002478:Cells, Cultured; D002642:Chick Embryo; D002851:Chromatography, High Pressure Liquid; D003094:Collagen / Q000096:biosynthesis*; D017364:Collagenases / Q000378:metabolism; D003470:Culture Media; D006133:Growth Substances / Q000494:pharmacology; D008751:Methylene Blue; D013194:Staining and Labeling; D014316:Tritium
D003470:Culture Media; D006133:Growth Substances; D014316:Tritium; D003094:Collagen; D017364:Collagenases; D008751:Methylene Blue