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Mouse Embryonic Stem Cell Adherent Cell Differentiation and Cytotoxicity (ACDC) Assay: Book Chapter
Citation:
BARRIER, M., K. J. CHANDLER, S. C. JEFFAY, M. R. HOOPES, T. B. KNUDSEN, AND E. S. HUNTER. Mouse Embryonic Stem Cell Adherent Cell Differentiation and Cytotoxicity (ACDC) Assay: Book Chapter. 2012 Edition, Chapter 11, Craig Harris; Jason M. Hansen (ed.), Developmental Toxicology; Methods and Protocols; Series: Methods in Molecular Biology, Springer Protocols. Humana Press Incorporated, Totowa, NJ, 889:181-196, (2012).
Impact/Purpose:
Embryonic stem cells have been used to evaluate the potential toxicity of chemicals in many studies. The adherent cell differentiation and cytotoxicity (ACDC) assay was developed using pluripotent mouse embryonic stem cells (mESCs) to evaluate chemical-induced effects on both stem cell viability and differentiation. This assay uses an In-Cell Western technique after a 9-day culture with DRAQ5/Sapphire700 cell/DNA stains used to quantify cell number and myosin heavy chain (MHC) protein is used as a marker of cardiomyocyte differentiation and is corrected for cell number, thereby separating cytotoxicity and effects on differentiation. 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, and has been applied to testing 309 unique environmental chemicals in the EPA’s ToxCast™ chemical prioritization research project. This chapter describes the experimental protocol for the ACDC assay.
Description:
There are thousands of environmental chemicals for which there is limited toxicological information, motivating the development and application of in vitro systems to profile the biological effects of xenobiotic exposure and predict their potential developmental hazard. An adherent cell differentiation and cytotoxicity (ACDC) assay was developed using pluripotent mouse embryonic stem cells (mESCs) to evaluate chemical-induced effects on both stem cell viability and differentiation. This assay uses an In-Cell Western technique after a 9-day culture with DRAQ5/Sapphire700 cell/DNA stains used to quantify cell number and myosin heavy chain (MHC) protein is used as a marker of cardiomyocyte differentiation and is corrected for cell number, thereby separating cytotoxicity and effects on differentiation. The ACDC assay can be used to evaluate the effects of xenobiotics on mESC differentiation and cell number in the same sample.
