Collagen is the major extracellular protein in mammals. Accurate quantification of collagen is essential in the biomaterials (e.g., reproducible collagen scaffold fabrication), drug discovery (e.g., assessment of collagen in pathophysiologies, such as fibrosis), and tissue engineering (e.g., quantification of cell-synthesized collagen) fields. Although measuring hydroxyproline content is the most widely used method to quantify collagen in biological specimens, the process is very laborious. To this end, the Sircol™ Collagen Assay is widely used due to its inherent simplicity and convenience. However, this method leads to overestimation of collagen content due to the interaction of Sirius red with basic amino acids of non-collagenous proteins. Herein, we describe the addition of an ultrafiltration purification step in the process to accurately determine collagen content in tissues.
Collagen quantification Colorimetric assay Biological specimens Ultrafiltration purification Sirius red
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This work forms part of the Teagasc Walsh Fellowship (Grant Award No. Ref: 2014045) and the ReValueProtein Research Project (Grant Award No. 11/F/043) supported by the Department of Agriculture, Food and the Marine (DAFM) under the National Development Plan 2007–2013 funded by the Irish Government. This work has also been supported from the Health Research Board, Health Research Awards Programme, under the grant agreement number HRA_POR/2011/84 and the Science Foundation Ireland and the European Regional Development Fund (Grant Agreement Number: 13/RC/2073).
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1.Regenerative, Modular & Developmental Engineering Laboratory (REMODEL)National University of Ireland GalwayGalwayIreland
2.Science Foundation Ireland Centre for Research in Medical Devices (CÚRAM)National University of Ireland GalwayGalwayIreland
3.Teagasc Research CentreDublinIreland
4.Centre for Cell Biology & Tissue Engineering, Competence Centre Tissue Engineering for Drug Development (TEDD), Department Life Sciences and Facility Management, Institute for Chemistry and Biotechnology (ICBT)Zürich University of Applied SciencesWädenswilSwitzerland