Collagen Boosting Test

Why conduct a collagen boosting test?

XCellR8’s in vitro collagen-boosting test (Sircol method) complies with our unique cruelty-free and vegan approach to testing. The primary application of the test is for efficacy testing and claim support in the cosmetics industry. In vitro mechanistic studies build confidence, helping to avoid lost time and revenue in the new product development (NPD) pipeline. The data can be used to support a variety of claims.

Here are some common applications of the method:

For ingredient suppliers:

• Independent data to demonstrate the collagen-boosting efficacy of ingredients.
• Scientifically robust, mechanistic evidence for use in case studies, brochures and presentations at conferences and trade shows.

For brands:

• Comparative screening of ingredients for efficacy and potency, for formulation development.
• Assessment of synergistic effects of actives.
• Screening blends or prototypes – confirming efficacy early in the NPD pipeline.
• Generation of in vitro data to build confidence as a prelude to clinical studies.
• Supplementation of clinical findings with mechanistic in vitro data, building a comprehensive support package for claims such as anti-wrinkle, anti-ageing, healthy ageing, collagen boosting, and improved skin texture.

Why animal-product-free?

The Sircol test aligns with XCellR8’s globally unique animal-product-free (APF) approach to in vitro testing. Most “non-animal” methods conducted globally still contain animal-derived components, such as foetal bovine serum (FBS), tissue extracts and antibodies. Key advantages of the APF approach include:

• Scientific: maximising the relevance of the test results for human physiology.
• Ethical: ensuring 100% animal-free testing for cruelty-free and vegan companies.

The Sircol method achieves Level 6 – within the green zone on XCellR8’s APF scale.

Overview of the test

Collagen is the major structural protein in human skin and is made by the dermal fibroblasts. Its key role in maintaining strength, firmness and elasticity makes collagen a key target for skincare products aimed at supporting healthy ageing.

The Sircol test provides robust in vitro data to demonstrate collagen boosting properties of cosmetic ingredients and formulations. Human dermal fibroblasts are exposed to the item for an agreed time (minimum 24 hours). Collagen is quantified using Sirius Red – a dye that binds with soluble collagen released into the culture medium. The results demonstrate the dose-dependent collagen-boosting potential of the test item.

Test system: human dermal fibroblasts in animal-product-free (APF) culture

The human dermal fibroblast cultures used in this test are obtained commercially as cryopreserved primary cells. They are originally derived from donor tissue (for example, following plastic surgery) with informed consent for the tissue to be used for research purposes, in compliance with the Human Tissue Act (UK) 2004. The cells have been extensively QC tested for a range of parameters including viability upon thawing from cryopreservation, proliferation rate, morphology and sterility (absence of bacteria, fungal growth and mycoplasma). They have also tested negative for HIV-1, HIV-2, HBV and HCV. They are maintained in the exponential growth phase in routine culture at 37°C / 5% CO2 prior to seeding into test plates. The culture medium and all culture reagents are free of animal-derived components, providing a fully human cell culture system.

SUMMARY OF THE TEST METHOD

Sircol is a colorimetric test for soluble collagen. Test items are assessed for solubility and compatibility prior to the study. Human dermal fibroblasts are seeded and collagen is quantified in the culture medium following an agreed exposure time (≥24 hours) with test item. The test contains a dye called Sirius Red that binds specifically with the Gly-X-Yn helical structure of soluble collagen, which is released into solution and can be measured by absorbance. Collagen of known concentrations can be used to generate a standard curve, and this can be used to calculate the concentration of collagen in test samples.

• Human dermal fibroblasts are seeded and allowed to adhere for 24 hours in a cell culture incubator at 37°C and 5% CO₂.
• Cells are exposed to fresh, serum-free media containing test item or vehicle control and cultures incubated for agreed exposure time (≥24 hours) in a cell culture incubator at 37°C and 5% CO₂.
• At the end of the exposure period, culture medium is collected and retained for analysis of collagen secretion.
• Note: culture medium can also be stored for further analysis of biomarkers (eg elastin) as a separate test if required. Please enquire for more details.
• Soluble collagen concentration is determined against a human placental collagen standard, using the Sircol colorimetric method.
• Collagen stain is applied to the samples and collagen-dye complexes precipitate out of solution.
• Dye is released from the complexes and solubilised, and the resulting red colour measured by absorbance using a spectrophotometer at a wavelength of 556nm.
• Data is expressed as a bar chart, with error bars and statistical significance, to demonstrate collagen boosting properties of the test item during the exposure period.

SAMPLE DATA

Figure 1 Sample data. A. Standard curve created using human placental collagen. B. Collagen release of HDF cells exposed to Test Item 1 at four concentrations for 24-hours (open bars) or 48-hours (solid bars). Bars represent mean collagen concentration calculated using the standard curve in A. Error bars represent the standard deviation. Statistical significance is calculated using Student’s two-tailed t-test. * p<0.05; ** p<0.005. VC = Vehicle Control. TC = Test Concentration.

Related Tests

Assessment of biomarkers associated with healthy ageing (eg elastin), using enzyme-linked immunosorbent assay (ELISA); changes in gene expression in the skin, using quantitative polymerase chain reaction (PCR); assessment of antioxidant potential, using the OxiSelect™ method.

Turnaround time

Standard 4-6 weeks. For urgent turnaround, please enquire.

Sample quantity

Standard request 10ml / 10g. We can work with less. Please enquire if supply is limited.

Pricing

Our test prices are dependent on the quantity of test items and any bespoke aspects of study design. Please contact us for a quote.