Limbal epithelial stem cell identification using immunoblotting analysis.

The unambiguous identification of limbal epithelial stem cells is currently a major challenge in corneal stem cell biology. Specific molecular markers which characterize these cells are lacking. At present, the best strategy for identification of limbal epithelial stem cells is to investigate a variety of putative markers for these cells in a differentiated (cytokeratin (CK) 3: CK3, integrin α6), undifferentiated (CK14), and naive state (∆Np63α, ATP-binding cassette subfamily G member 2 (ABCG2), integrin α9, Notch-1), alongside functional assays which indicate their stemness. The focus of this chapter is to highlight advances in the Western blotting technique for quantitative assessment of corneal epithelial cell markers, and the use of this technique for investigation of a range of different protein markers which identify limbal epithelial stem cells.