In Vitro Micronucleus Test

According to OECD Guidelines for Testing of Chemicals, Test No. 487, the in vitro micronucleus (MNvit) test is used to evaluate the presence of micronuclei (MN) in the cytoplasm of interphase cells.

STEMart provides in vitro micronucleus test to determine whether a medical device is genotoxic to cells in culture. This assay is available under either GLP (Good Laboratory Practice) or Non-GLP conditions.

Standard

OECD 487

Test Cell

• Human peripheral blood lymphocytes (HPBLs)
• Chinese Hamster Ovary cell line (CHO-K1)
• Chinese Hamster Lung cell line (CHL, V79)
• TK6 Human Lymphoblastoid cells
• L5178Y

Test Method

Fig. 1 In vitro micronucleus test procedure. (Migliore, L. 2014)

Cells are incubated in 96-well plate with addition of at least three concentrations with duplicates of the medical device extract for a short period (3 to 6 hours) in the presence and absence of metabolic activation (S9) and for a long period in the absence of S9. (CytoB is used as a cytokinesis blocker when human lymphocytes are used.)

Cells are incubated on plate for 1.5-2 cell cycle lengths following the end of treatment to allow chromosome damage on cell cycle/cell division to lead to the formation of micronuclei.

Chromosomal spread slides are then prepared and stained with Giemsa, fluorescent DNA specific dyes or other methods. The presence of micronucleus is evaluated under microscopy in at least 4000 bi-nucleated cells from each concentration (2000 cells from each of the duplicates).

Flow cytometry and laser scanning cytometry can be used for cell analysis.

Negative vehicle control and positive controls are included in each study.

Criteria

• Positive result: A concentration-related increase or a reproducible increase in the percentage of cells with micronuclei over that of the solvent controls.
• Negative result: No statistically significant increase in the percentage of cells with micronuclei.

Final Report

The final report including information on the methodology, raw data, analysis, and interpretation of the results will be provided for customer.

Advantages

• Comprehensively investigate chromosome damaging potential of both aneugens and clastogens, which can induce chromosome loss and chromosome breaks respectively, in a single endpoint.
• The combination of telomer and centromer staining increases sensitivity and allow discrimination between aneugens and clastogens.
• The assessment of micronuclei can be scored faster because the in vitro micronucleus test examines cells at interphase.
• Automated scoring is available with flow cytometry.

If you have additional questions about in vitro micronucleus test, or would like to find out more about our services, please feel free to contact us.

Reference

• Migliore, L., et al. "The in vitro micronucleus assay and FISH analysis." Genotoxicity and DNA Repair. Humana Press, New York, NY, 73-102 (2014).
• OECD Guideline for Testing of Chemicals. No 487 In Vitro Mammalian Cell Micronucleus Test (2014).