PCNA immunostaining

Immunostaining staining was carried out using primary antiserum to proliferating cell nuclear antigen (PCNA) (PC10, Santa Cruz Biotechnology Inc., Heidelberg, Germany). Briefly, the primary antibody was diluted in Tris buffer with a dilution of 1:50 (as determined by the data sheet). The sections were incubated with the primary antibody overnight at + 4 °C. The binding of the primary antibody was observed using a commercial avidinbiotin- peroxidase detection system recommended by the manufacturer (DAKO, Carpenteria, USA). Finally, the slides were stained with diaminobenzene (DAB).