I have been informed that there is a discrepancy between the OECD TG442F on the
h-CLAT assay and the DB-ALM protocol. The test guideline cannot be officially
updated until summer 2017; until then please share this attached, corrected
Propidium iodide (PI) staining
25. After 24±0.5 hours of exposure, cells are transferred into sample tubes and
collected by centrifugation.
The supernatants are discarded and the remaining cells are resuspended with 200
μL (in case of 96-well) or
600 μL (in case of 24-well) of a phosphate buffered saline containing 0.1%
bovine serum albumin (staining
buffer). 200 μL of cell suspension is transferred into 96-well round-bottom
plate (in case of 96-well) or
micro tube (in case of 24-well) and washed twice with 200 μL (in case of
96-well) or 600 μL (in case of 24-
well) of staining buffer. Finally, cells are resuspended in staining buffer
(e.g. 400 μL) and PI solution (e.g.
20 μL) is added (for example, final concentration of PI is 0.625 μg/mL).
Description: TG 442E_ENG_REV2017.pdf