In order to standardize the implementation of biocompatibility evaluation standards such as GB/T 16886 series and facilitate the preparation of relevant chapters of registered product standards, the biocompatibility laboratory of our institute has prepared examples of biocompatibility test project requirements and test methods for the reference of relevant personnel.

Biocompatibility test requirements

Common test items:

1. Cytotoxicity: The relative cell proliferation rate should not be less than 70%. (When using MTT method in GB/T16886.5-2017) the cytotoxicity should not be greater than grade X. (The requirements of AGAR diffusion method, etc., the requirements of enterprises to determine their level according to the nature and risk of their own products)

2. Intradermal reaction: The average score difference between the test sample and the solvent control should not be greater than 1.0.

3. Delayed hypersensitivity: There should be no delayed hypersensitivity.

Other test items

Classification of stimulation tests:

4. Primary skin irritation: Primary skin irritation index should not be greater than 0.4.

5. Vaginal stimulation: The stimulation index should not be greater than 4 (very light) /8 (mild). (Enterprises according to their own product nature and risk requirements)

6. Oral mucosal irritation: The stimulation index should not be greater than 4 (very light) /8 (mild). (Enterprises according to their own product nature and risk requirements)

7. Eye irritation: The test sample does not cause eye irritation.

Systemic Toxicity test classification:

8. Acute systemic toxicity: There should be no acute systemic toxicity.

9. Subacute systemic toxicity: There should be no subacute systemic toxicity.

10. Subchronic systemic toxicity: there should be no subchronic systemic toxicity.

11. Pyrogen: The test sample should have no pyrogen reaction.

Genotoxicity test:

12. Salmonella Typhimurium back mutation test (Ames test) : The mutagenesis should be negative.

13. Mouse lymphoma cell gene mutation test: mutagenesis should be negative.

14. Chromosome aberration test: mutagenesis should be negative.

15. Implantation tests: should cause no or minor histological reactions compared to control samples. (Note: The implantation test should specify the implantation site and period, such as 12 weeks muscle implantation test)

Biocompatibility test method:

Principle: Mainly define the extraction medium, extraction proportion, extraction temperature, extraction time and the method used

1. Cytotoxicity: Complete cell medium was used as the extraction medium, and was extracted at (37±1) ℃ at a ratio of 0.2g/mL for 24 h, according to the extraction method specified in GB/T 16886.5-2017. The results should meet the requirements of X.X.

Note: The extraction ratio shall be selected by the enterprise according to the characteristics of the sample according to the corresponding conditions in GB/T 16886.12; At the same time, according to the selection of different cytotoxicity test methods (such as extract method MTT, AGAR diffusion method, direct contact method, etc.) are described in the method.

2. Intradermal reaction: Normal saline and vegetable oil were used as the extraction medium respectively, at the ratio of 0.2g/mL, at the condition of (37±1) ℃ for (72±2) h, according to the intradermal reaction method specified in GB/T 16886.10-2017. The results should meet the requirements of X.X.

Note: If you choose primary skin stimulation, vaginal stimulation, oral mucosal stimulation, eye stimulation, please replace the intradermal response method at the corresponding position of the standard; At the same time, contact the engineer to determine the type of extraction medium used, or whether the sample is directly tested as the test object.

3. Delayed hypersensitivity reaction: Normal saline and vegetable oil were used as the extraction medium respectively, at the ratio of 0.2g/mL, at the condition of (37±1) ℃ for (72±2) h, according to the maximum dose method specified in GB/T 16886.10-2017, and the results should meet the requirements of X.X.

Note: According to the nature of the sample tested, the enterprise chooses the maximum dose method or the paste method.

4. Acute systemic toxicity: Normal saline and vegetable oil were used as the extraction medium, at the ratio of 0.2g/mL, at the condition of (37±1) ℃ for (72±2) h, and according to the methods of caudal vein and intrabitoneal injection specified in GB/T 16886.11-2011, the test results should meet the requirements of X.X.

Note: According to the application path of the product, the enterprise chooses the corresponding drug delivery route, such as the oral route, and replaces it in the corresponding position.

5. Subacute systemic toxicity: Normal saline was used as the extraction medium, at the ratio of 0.2g/mL, at the condition of (37±1) ℃ for (72±2) h, and the test was conducted according to the method of continuous injection into the tail vein for 14 days (or oral gavage for 28 days) prescribed in GB/T 16886.11-2011. The results should meet the requirements of X.X.

6. Subchronic systemic toxicity: Normal saline was used as the extraction medium, at the ratio of 0.2g/mL, at the condition of (37±1) ℃ for (72±2) h, and the test was conducted according to the method of continuous injection into the tail vein for 28 days (or oral gavage for 90 days) prescribed in GB/T 16886.11-2011. The results should meet the requirements of X.X.

Note: According to the characteristics of the product and the way of application to choose the route of administration (intravenous or oral)

7. Pyrogen: Normal saline was used as the extraction medium, at the ratio of 0.2g/mL, at the condition of (37±1) ℃ for (72±2) h, and the test was carried out according to the method prescribed by the Chinese Pharmacopoeia. The results should meet the requirements of X.X.

8. Salmonella Typhimurium return mutation test (Ames test) : Normal saline and DMSO were used as the extraction medium, respectively, at the ratio of 0.2g/mL at (37±1) ℃ for 72 h, according to the method specified in GB/T 16886.3-2008, the test results should meet the requirements of X.X.

9. Mouse lymphoma gene mutation test: Normal saline and DMSO were used as the extraction medium respectively, at the ratio of 0.2g/mL, at (37±1) ℃ for 72 h, according to the method specified in GB/T 16886.3-2008, the test results should meet the requirements of X.X.

10. Chromosome aberration test: Normal saline and DMSO were used as the extraction medium, respectively, at the ratio of 0.2g/mL, at the condition of (37±1) ℃ for 72 h, according to the method specified in GB/T 16886.3-2008, the test results should meet the requirements of X.X.

11. Implantation test: The sample is prepared into a suitable size of the subject, and implanted into the corresponding part of the experimental animal by subcutaneous/muscle/bone method, the implantation cycle is X weeks, and the test is conducted according to the corresponding method specified in GB/T 16886.6-2015, and the results should meet the requirements of X.X.

Source: Medical Device Technical Evaluation Center, State Drug Administration