In addition to the capability of the Comet assay to identify DNA damage at the single cell level, other significant advantages include its sensitivity for detecting low levels of DNA damage, the requirement for only small numbers of cells per sample, its ease of application, and the short time needed to perform the assay.
2Liao W, McNutt MA and Zhu W-G (2009) Methods 48; 46-53
|Test System||Alkaline Comet (single cell gel electrophoresis)|
|Cell Line||TK6 cells|
|Test Article Concentration||5 serial dilutions, typically 2-fold; e.g. 1000, 500, 250, 125, 62.5 µg/mL|
|Incubation Time||3 hr exposure time|
|Quality Controls||1% DMSO (negative control)
Etoposide (positive genotoxic control)
|Cell Viability Assesment||Propidium iodide exclusion using flow cytometry; 80% relative survival required for Comet processing|
|Test Article Requirements||10 mg solid - Other compound preparations are acceptable|
|Data Analysis||50 comets per compound dose acquired from two agarose gels (25 from each gel)|
|Data Delivery||Written report presenting overall results
Excel worksheets with full graphical dose response data for genotoxicity including % tail intensity
The in vitro Comet assay from Gentronix has been validated against a number of different genotoxic and non-genotoxic compounds.
1 Singh NP et al. (1988) Exp Cell Res 175; 184-191
2 Liao W et al. (2009) Methods 48; 46-53