Soluble Collagen Assay Kit (ab241015)
Key features and details
- Detection method: Fluorescent
Overview
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Product name
Soluble Collagen Assay Kit
See all Collagen kits -
Detection method
Fluorescent -
Product overview
Soluble Collagen Assay Kit (ab241015) detects newly synthesized (acid-soluble) collagen levels in various biological samples and cultured cell medium.
The assay involves extraction of soluble collagen in acetic acid, followed by enzymatic degradation of collagen into glycine-rich oligopeptides, which are quantified using a fluorogenic reagent and developer solution that selectively react with the N-terminal glycine fragments to form a stable fluorescent complex (Ex/Em = 376/468 nm).
The assay is more sensitive and selective than dye-binding (Picrosirius Red) soluble collagen assays, is simple to perform and has a linear range from 0.05 – 2 μg collagen per well (2.5 μg/mL to 100 μg/mL in a 20 μL sample volume).
Properties
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Storage instructions
Store at -20°C. Please refer to protocols. -
Components 100 tests Collagen Assay Buffer 1 x 25ml Collagen I Standard (2 mg/ml) 1 x 100µl Collagenase Enzyme Mix 1 vial Detection Reaction Buffer 1 x 10ml Developer Solution 1 x 500µl Peptide Labelling Reagent 1 vial
Images
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Collagen I Standard curve.Collagen I Standard curve.
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Estimation of acid-soluble collagen content in rat tissues.Rat leg muscle, lung and heart samples were homogenized in 0.5 M acetic acid, incubated overnight at 4C to solubilize collagen and neutralized with 0.5 M NaOH. Collagen levels (calculated as µg collagen/mg wet tissue) for the samples were: 3.83 ± 0.47 µg/mg for muscle, 1.47 ± 0.22 µg/mg for lung and 2.85 ± 0.08 µg/mg for heart.
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Estimation of secreted soluble collagen in cultured cell growth medium (DMEM/F12 medium).3T3-L1 fibroblasts were grown to ~80% confluence in T-75 flasks and then treated with either vehicle or TGF-β (100 ng/ml), a cytokine known to stimulate synthesis and secretion of tropocollagen fibrils. After 48 hours of treatment, the culture medium was removed, centrifuged to remove debris and was assayed directly (each 10 µl clarified medium, undiluted). TGF-β treatment resulted in a roughly 1.5- fold increase in collagen secretion. Data are mean ± SEM of 3 replicates, assayed according to the kit protocol.
