Figure 8.

Clonal analysis of cells isolated from limbal epithelium and stroma. Cells from the surface epithelium (isolated by dispase) or the remaining stroma (isolated by collagenase) were seeded on 3T3 fibroblast feeder layers. Epithelial colonies were readily detected by crystal violet staining from cells of isolated surface epithelium of the normal control, AL explant, and submerged explant (A, left). Nevertheless, epithelial colonies were not formed by cells isolated from the stroma of the normal control and submerged explants, but formed by AL explants (A, right). B: Epithelial colonies were stained dark with a smooth and distinct border as verified by higher magnification. C: In contrast, fibroblastic colonies were stained faint with an irregular border as verified by higher magnification. Those stromal colonies of AL (B) have clearly p63-positive expression in all of cells (D), and elongated fibroblast-like p63-positive cells were observed out of colony (D, inset). E: These colonies expressed pancytokeratins by AE1/AE3. Scale bars, 50 μm.