Immunohistochemical detection of proliferating lipocytes in regenerating rat liver

For the detection of proliferating lipocytes in regenerating liver, partially hepatectomized rats were injected intraperitoneally with 5-bromo-2'-deoxyuridine (BrdUrd, 50 mg/kg body weight) and killed 1 h later. Acetone-fixed frozen liver sections were used for the simultaneous detection of cytoplasmic desmin and BrdUrd-labelled nuclei of lipocytes using double immunohistochemical procedures. The best results were obtained with the sequences: rabbit anti-desmin----biotinylated anti-rabbit IgG----avidin-biotin-peroxidase complex----3,3'-diaminobenzidine tetrahydrochloride, followed by DNA denaturation----mouse anti-BrdUrd----anti-mouse IgG----peroxidase-anti-peroxidase complex----4-chloro-1-naphthol. With this method, cytoplasmic desmin was stained a brown colour, which sharply contrasted with the blue-stained BrdUrd-labelled nuclei. Unlabelled nuclei appeared green after counterstaining with methyl green. No cross-reaction between immunoreagents of desmin and BrdUrd stainings was observed. The labelling index of lipocytes peaked (25.7 per cent) 48 h after partial hepatectomy, whereas it was 3.7 per cent in normal rat liver.