Quantitative analyses by radioimmunoassay of proliferating cell nuclear antigen (PCNA/cyclin) in ocular tissues

A radioimmunoassay (RIA) was developed for a proliferating cell nuclear antigen, PCNA/cyclin, which is a protein synthesized in phase with DNA as a marker for cell replication. This antigen was used in rats to detect amounts of PCNA and to estimate rates of cell replication in ocular tissues (cornea, lens epithelium, retina-choroid and optic nerve) and in nonocular tissues (skin, skeletal muscle, brain, thymus, small intestine and liver). Antisera were raised in rabbits to the oligopeptide fragment of human PCNA (hPCNA): the C-terminal fragment [Tyr254]-hPCNA (254-261). For the RIA, synthetic peptides were used as the standard, and the radioiodinated [125I-Tyr254]-hPCNA (254-261) was used as the radioligand. The IC50 was 30-40 fmole/100 microliters with a sensitivity of 0.5-1.0 fmole/100 microliters for this assay. Tissue extracts in 1 N acetic acid were digested with trypsin to release the fragment (254-261) from whole PCNA for assay. The highest amounts of immunoreactive PCNA were found in lens epithelium and skin, while the lowest amounts were found in optic nerve and muscle. The cornea showed a higher value than the retina-plus-choroid or the optic nerve. These results suggest a high rate of cell replication in lens epithelium.