A simple and sensitive method for the quantitative estimation of collagen

doi:10.1016/0003-2697(79)90795-4

Analytical Biochemistry

Volume 94, Issue 1, 1 April 1979, Pages 96-99

https://doi.org/10.1016/0003-2697(79)90795-4 Get rights and content

Abstract

A method to quantitate collagen in solution is described. It is based on binding of the dye Sirius Supra red F3BA by collagen followed by elution and estimation of the bound dye in a spectrophotometer. The assay can be used in the range from 10 to 100 μg protein. The method is rapid, specific, simple, sensitive, and highly reproducible.

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Metals and their metal ions have been shown to exhibit certain biological functions that make them attractive for use in biomaterials, for example in bone tissue engineering (BTE) applications. Recent data shows that Molybdenum (Mo) is a potent inducer of osteogenic differentiation in human bone marrow-derived mesenchymal stromal cells (BMSCs). On the other hand, while boron (B) has been shown to enhance vascularization in BTE applications, its impact on osteogenic differentiation is volatile: while improved osteogenic differentiation has been described, other data show that B might slow down osteogenic differentiation or reduce the calcification of the extracellular matrix (ECM) when applied in higher doses. Still, the combination of pro-osteogenic Mo and pro-angiogenic B is certainly attractive in the context of biomaterials intended for the use in BTE.
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Molybdenum trioxide enhances viability, osteogenic differentiation and extracellular matrix formation of human bone marrow-derived mesenchymal stromal cells
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Citation Excerpt :
Expression of COL1A1, a relevant marker gene for ECM formation [40], was diminished in a concentration-dependent manner in MoO3 groups on D7 and only a slight upregulation above the control’s level was observed thereafter. Nevertheless, the actual amount of collagen produced and deposited by BMSCs, measured via Sirius Red staining [29–31], was found to be significantly increased in presence of 1 and 2 mM MoO3. Possible explanations for the distinct differences between gene expression levels and the proteins’ actual production are discussed controversially.
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Citation Excerpt :
The absorbance of the elution was detected at 650 nm using a microplate reader (Vmax, Molecular Devices, CA, USA). Collagen fibers were detected by the SR method [27]. The cells were fixed with 4% paraformaldehyde for 30 min at 4 °C and stained with 0.1% Sirius Red F3B in a saturated aqueous solution of picric acid for 1 h.
Isoflavones are phytoestrogens abundant in leguminous crops and are used to prevent a variety of hormonal disorders. In the present study, the effects of genistein and daidzein on the chondrogenic differentiation of ATDC5 cells were investigated. Genistein (10 μM) treatment markedly reduced production of sulfated proteoglycans and collagen fibers in the ATDC5 cells. Genistein suppressed the expression of genes involved in chondrocyte differentiation such as Sox9, Col2a1, Col10a1, Acan, and Tgfb1. Additionally, genistein significantly decreased calcium deposition in ATDC5 cells during chondrogenic differentiation; however, it increased calcification under non-chondrogenic mineralizing conditions. Daidzein exhibited a similar effect of suppressing chondrogenesis in ATDC5 cells, although its efficacy was 10-times lower than that of genistein. These findings suggest that a high concentration of genistein inhibits chondrogenesis and chondrogenic mineralization, whereas it enhances non-chondrogenic mineralization.
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Oxidative stress, the disrupted oxidation-reduction mechanism in our body, is caused by the excessive exposure of free radicals and the impaired antioxidant defenses that can accelerate skin aging. Antioxidants can be obtained from nature, which are available widely in therapeutic-rich plants, such as white saffron (Curcuma mangga Val., denoted as C. mangga). Although many pieces of evidence reveal that C. mangga contains an abundance of phenolic compounds and has antioxidative effects, its cosmeceutical potentials remain unclear. The present study aimed to disclose the unexplored antiaging potentials of C. mangga extract (CME) in oxidative stress-induced human BJ fibroblasts with a focus on collagen protection against pro-inflammatory mediators MMP1, MMP3, and MMP13. The oxidative stress-induced cells were treated with CME and curcumin at different doses. The results showed that treatment using CME (25 μg/mL) could maintain the collagen contents up to 18.45 ± 0.68 μg/mL in H₂O₂-treated fibroblasts (only ~26.63% reduction in collagen contents), while the figure for the negative control was the lowest (12.79 μg/mL), showing a significant reduction in collagen contents by 49.13%. In addition, the gene expression of pro-inflammatory MMPs arose significantly in BJ fibroblasts after oxidative stress induction using 200 μM H₂O₂, in which the expression for MMP1, MMP3, and MMP13 increased by 7.10, 38.96, and 2.69 times, respectively. Interestingly, CME treatment (100 μg/mL) could effectively inhibit MMP1, MMP3, and MMP13 gene expression by 3.65, 34.62, and 2.02 times, respectively. In conclusion, CME showed favorable antiaging activities in H₂O₂-treated human BJ fibroblasts as confirmed by the low levels of gene expression of MPP1, MMP3, and MMP13 after treatment with CME.
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A simple and sensitive method for the quantitative estimation of collagen

Abstract

Anal. Biochem

Arch. Pathol