The use of novel antibodies to identify substates within the stem cell compartment of human ES cells

Various recent data point to the heterogeneity of undifferentiated stem cells, which appear to be able to exist in alternative states that can interconvert from one to another. The precise significance of these ‘sub-states’ remains to be established. It may be that they may represent different steps as a stem cell prepares to differentiate so that, while not committed to differentiate, cells in some states are more likely to differentiate than those in other states. Another possibility is that the sub-states exhibit lineage priming so that cells in one sub-state might be more likely to differentiate into a particular lineage than cells in another sub-state at the time when they commit to differentiate. To test these hypotheses it is essential to be able to identify and isolate cells in particular sub-states. The phenomenon of ‘culture adaptation’ of human ES cells tends to ‘trap’ cells in sub-states in a way that permits their isolation. Otherwise, prior to culture adaptation it is thought that some such sub-states may exist but only in a transitory way, making it difficult to isolate them. Consequently in this project we are utilising the EC cell line NTERA2, which given its oncogenic nature it can be regarded as a culture adapted cell line. We are also employing the well characterised, culture adapted, human ES cell line, H7.S6, together with several newly derived antibodies that mark the stem cell state, in conjunction with other established markers, to define, isolate and characterise sub-states that may exist within the undifferentiated stem cell compartment of human ES cells.