Expansion of the Embryonic Stem Cell Test: Differentiation into neural cells
2005
Hayess,K. | Buesen,R. | Schlechter,K. | Slawik,B. | Spielmann,H. | Seiler,A.
Alemán. The embryonic stem cell test (EST) is a validated in vitro assay that has been established to classify compounds with respect to their embryotoxic potential. The current experimental procedure involves differentiation of murine embryonic stem cells (D3) into contracting cardiomyocytes. However, potentially embryotoxic drugs may effect primarily other tissues than myocard. Consequently, this consideration prompted us to expand the EST to other major target tissues.Here, we present a protocol for differentiation of murine embryonic stem cells into neurons designed with special regard to the testing of chemicals. This modified protocol is based on a monolayer differentiation procedure and offers the advantage of a reproducible development of neural cells in a comparatively short time. The differentiation of D3 cells into neural cells was characterised by analysis of neuron-specific marker gene expression using flow cytometry. In addition, the developing neurons were examined by immunofluorescence staining using neuron-specific antibodies. As a result, we were able to define neuron-specific molecular endpoints for the detection of chemical effects on embryonic development.The expansion of the EST to more than one target tissue will considerably improve the accuracy of this predictive screen by preventing false negative results.
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