Isolation and characterization of a mouse monoclonal antibody against human corneal endothelial cells

In vitro cultivation of human corneal endothelial cells (HCEC) is associate d with loss of typical cobblestone-like appearance during successive passag es. Thus far morphology was the sole criterion for the cell's endothelial n ature. Mouse monoclonal antibodies (mabs) to human corneal endothelial cell s were raised using standard immunization and hybridoma isolation procedure s. The specificity of mabs fbr human corneal endothelial cells was tested i n comparison to other endothelial cell types, to fibroblasts, corneal kerat ocytes and to human retinal pigmented epithelial cells. In addition immunof luorescence or immunoperoxidase staining was performed with frozen tissue s ections of human corneas and with various other human tissues. The mab 9.3. E reacts with cultured human corneal endothelial cells, but not with cultur ed human fibroblasts and human keratocytes. In frozen sections selective po sitivity of corneal endothelium in contrast to negativity of the other corn eal cell types was confirmed. In investigated extraocular tissues positivit y was observed in smooth muscle cells including related cells (i.e. Ito and mesangial cells) and in Schwann's cells and adipocytes, but apparently not in vascular endothelial cells. The mab is human-specific and binds to a pr otein with a molecular weight of 130 kDa mainly accumulating along cell mem branes. A mouse monoclonal antibody against human corneal endothelial cells was established in vitro and was shown to be capable of differentiating co rneal endothelial cells from other corneal cell types. especially from corn eal keratocytes. It is, however, not cornea-specific, but also reacts with certain extraocular cell types. (C) 2001 Academic Press.