Mouse embryonic stem cell adherent cell differentiation and cytotoxicity (ACDC) assay

An adherent cell differentiation and cytotoxicity (ACDC) assay was developed using pluripotent J1 mouse embryonic stem cells (mESCs). Adherent mESCs were used to evaluate chemical-induced effects on both stem cell viability and differentiation using an in-cell western technique after a 9-day culture. DRAQ5/Sapphire700 stains were used to quantify cell number. Myosin heavy chain protein was used as a marker of cardiomyocyte differentiation and was corrected for cell number, thereby separating cytotoxicity and effects on differentiation. Acetic acid, 5-fluorouracil and bromochloroacetic acid were evaluated using the embryonic stem cell test and ACDC assay. Both systems distinguish the relative potencies of these compounds. TaqMan low-density arrays were used to characterize the time course of differentiation and effects of chemical exposure on multiple differentiation gene markers. The ACDC assay is a technique that can be used to evaluate the effects of xenobiotics on mESC differentiation and cell number using a single assay.