Measuring soluble collagen in biological samples is a straightforward experiment which can be done with the Sircol soluble collagen assay using Sirius Red dye (Sircol dye) in a convenient manner (we explored this method in a recent post). Nevertheless, up to now it has been a challenge to measure insoluble collagen.
During periods of rapid growth, development, tissue repair remodeling and wound healing, insoluble collagen fibers are laid down. Until now, an investigator seeking to measure covalent cross linked insoluble collagen required measurements based on free hydroxyproline content, a procedure that requires strong acid (6.0 M HCl), high temperatures (+ 950 C) and overnight cooking (18 to 24 hours).
Insoluble collagen measurements made easy
By applying both assays, the Sircol Collagen assay and the Sircol INSOLUBLE Collagen Assay, even three collagen fractions can be differentiated:- Collagen soluble in cold dilute acid
- Collagen soluble in cold dilute acid and pepsin
- Insoluble collagen
Of course, by adding the results together a value for the “Total Collagen” in a sample can be established.
Interested in using Sircol assays or outsourcing them?
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6 Responses
Dear Dr. El-Baya,
Do you have experience with measuring of collagens (soluble and insoluble fractions) in blood of blood-based fluids (serum, plasma)? Can you distinguish which collagens you measure?
Many thanks!
Best regards,
Helge Taubert PhD
Dear Dr. Taubert,
as the kits to measure the insoluble fraction of collagen are quite new on the market, I have to check with the laboratory. I will come back to you asap.
Best
Ali
Dear Dr. Taubert and all others who might be interested in the question whether the Sircol collagen assay can be used with blood (or serum/plasma) samples.
Unfortunately this application is not recommended due to the high albumin interference.
Best
Ali
Dear Dr. el Baya,
Many thanks for your efforts and answer! It helps us not to perform unsuccessful experiments.
Best regards,
Helge Taubert
Can your kit measure soluble human collagen in cell culture medium? The medium has Fetal bovine serum at 10% in it.
Dear Jeffrey,
Thank you for your question on our blog.
You can indeed measure soluble collagen with our SIRCOL assay.
Where the collagen concentration is less than 5µg/100µl, a Collagen Isolation & Concentration step is recommended.
This step requires overnight incubation at 4oC. Low protein binding microcentrifuge tubes should be used when the culture medium contains a serum supplement.
See page 5 of manual at this link: http://www.biocolor.co.uk/site/wp-content/uploads/2016/04/sircol-soluble-assay.pdf
I hope this helps.
Best of luck with your project!
Isabelle