Immunohistochemical demonstration of proliferating cell nuclear antigen(PCNA) in formalin-fixed. paraffin-embedded sections from rat and human tissues.

Abstract

Proliferating cell nuclear antigen (PCNA) was demonstrated in paraffin-embedded tissues by an indirect immunohistochemical method using both IgG₁ (19F4) and IgM (19A2) class monoclonal antibodies (MAbs). Various fixatives were compared, at different temperatures and different fixation times, and pretreatment procedures prior to staining were also studied. PCNA was demonstrated when tissues were fixed in aldehyde fixative for a short time; the staining was more intense when tissues were fixed at 4°C than when they were fixed at room temperature. Longer fixation times were associated with less intense staining, and none of the pretreatments tested prevented the loss of immunohistochemical reactivity in such cases. Sufficient PCNA staining was obtained in methanol-fixed tissues; the staining was reduced when tissues were fixed in ethanol. This investigation showed that the usual 10% formalin-fixed paraffinembedded tissues were applicable for PCNA immunohistochemistry when the fixation time was less than 24 hr, even at room temperature. Under these conditions, PCNA-positive nuclei were clearly demonstrated in physiologically growing and cancerous rat and human tissues.