Cytotoxicity Testing

The ability of a substance to damage cells or tissue is referred to as cytotoxicity.

Cytotoxicity tests represent one of the easiest methods within the scope of the biological evaluation of medical devices (standard series DIN EN ISO 10993-5 and 10993-12) to analyse the detrimental effects of substances. Cell culture techniques allow a rapid yet sensitive diagnosis of the biological reactivity of leachable or diffusible components of materials.

The safety of the medical devices which are used in or on human beings is the main concern of this evaluation. Testing methods using cell cultures have shown good correlation with animal assays in many cases.

Which test system is the best for my product?

• BCA can be used for submissions in Central Europe, while XTT and MTT both can be used for submissions worldwide. All three methods are performed with extracts or solutions of the test material (ISO 10993-5). The XTT assay is preferred over the MTT assay as it produces a water soluble formazan, thus not requiring a solubilisation step before measuring.
• The Elution Test is designed for the evaluation of extracts of elastomeric plastics or polymeric materials. It is appropriate for high-density materials (U.S. Pharmacopeia <87>). This test system can be combined with ISO for dose-response evaluations and quantitative assessment of cytotoxicity (ISO 10993-5).
• The Colony Forming Assay is specifically required by the authorities in Japan (ISO 10993-5, MHLW-guideline).
• The Direct Cell Contact Test is designed for test materials with a flat surface that are neither low nor high-density materials. The test item is applied directly to the test system (ISO 10993-5).
• The Agar Diffusion Test is an option if the test item cannot be extracted or dissolved in cell culture medium. It is also relevant that the test item does diffuse through, but does not react with the agar layer. The sponsor has to provide a justification for the test method in order to submit his test material with this test method (ISO 10993-5, ISO 7405).

References

1. ISO 10993-5: 2009, “Biological evaluation of medical devices – Part 5: Tests for in vitro cytotoxicity“
2. ISO 10993-12: 2012, “Biological evaluation of medical devices – Part 12: Sample preparation and reference materials“
3. United States Pharmacopeia and National Formulary (USP 41), Chapter <87> (2018). Biological Reactivity Tests, in vitro - Elution Test
4. ISO 7405: 2018, “Dentistry - Evaluation of biocompatibility of medical devices used in dentistry - Test procedures specific to dental materials”
5. MHLW (Ministry of Health, Labour and Welfare, Japan) Notification by Director of OMDE Yakushokuki-hatsu No. 0301-20, “Basic Principles of Biological Safety Evaluation Required for Application for Approval to Manufacture/Market Medical Devices”, March 1, 2012
6. MHLW (Ministry of Health, Labour and Welfare, Japan) Ministerial Notification by Director of Health, Labour and Welfare Yasuhisa Shiozaki No. 64, The Japanese Pharmacopoeia, Seventeenth Edition, March 07, 2016, chap.7.03, 4. “Cytotoxictity test”