Essential modification of the Sircol Collagen Assay for the accurate quantification of collagen content in complex protein solutions

被引：57

作者：

Lareu, Ricky R. ^{[2
,3
]}

Zeugolis, Dimitrios I. ^{[1
,3
,4
]}

Abu-Rub, Mohammad ^{[1
]}

Pandit, Abhay ^{[1
]}

Raghunath, Michael ^{[3
,4
]}

机构：

[1] Natl Univ Ireland Galway NUIG, Network Excellence Funct Biomat NFB, Galway, Ireland

[2] Natl Univ Singapore, Yong Loo Lin Sch Med, Dept Orthoped Surg, NUSTEP, Singapore 117595, Singapore

[3] Natl Univ Singapore, Fac Engn, Div Bioengn, Tissue Modulat Lab, Singapore 117548, Singapore

[4] Natl Univ Singapore, Yong Loo Lin Sch Med, Dept Biochem, Singapore 117595, Singapore

来源：

ACTA BIOMATERIALIA | 2010年 / 6卷 / 08期

基金：

爱尔兰科学基金会;

关键词：

Hydroxyproline assay; Sircol assay; Collagen content; Sirius red; Ultrafiltration; RED F3BA; FIBERS; TISSUE; CELLS; HYDROXYPROLINE; CHONDROCYTES; QUANTITATION; MICROSCOPY; CULTURE; ABSENCE;

D O I：

10.1016/j.actbio.2010.02.004

中图分类号：

R318 [生物医学工程];

学科分类号：

0831 ;

摘要：

Collagen contains the unique imino acid hydroxyproline (HyPro). which is involved in the stabilization of this triple helical molecule The concentration of HyPro is customarily used to calculate the total collagen content in a cell culture environment and in acid hydrolysates of normal and pathophysiological tissues. Radiolabelling, chromatographic and calorimetric assays have been developed over the years for the accurate determination of collagen content through HyPro estimation Recently, the Sircol Collagen Assay (SCA) has been almost exclusively adopted as the fastest and simplest colorimetric method for the determination of collagen concentration in complex protein solutions We show here that the colorimetric SCA, which is based on the binding of Sinus red (SR) to collagen, is flawed by interference of non-collagenous proteins (e g scrum) In fact, we demonstrate that SCA in cell culture systems and tissue hydrolysates results in a dramatic overestimation of collagen content ranging from 3- to 24-fold In order to rescue this otherwise very practical assay, we introduce a simple purification procedure that allows the removal of interfering non-collagenous proteins from culture media and tissue samples so that accurate measurements with SCA are now possible (C) 2010 Acta Materialia Inc. Published by Elsevier Ltd All rights reserved

引用

页码：3146 / 3151

页数：6

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