This study uses CD10 and epithelial membrane antigen (EMA) as respective proximal and distal tubular segment markers to localize polyoma virus replicative activity, as determined by large T antigen (TAg) expression, in allograft polyoma virus nephropathy (PVN). Sixteen biopsies and 2 nephrectomy specimens with PVN had serial 2-mum paraffin sections stained using monoclonal antibodies to polyoma virus TAg by immunoperoxidase with diaminobenzidine as chromogen. A second immunolabeling step used CD10 as a proximal nephron marker or EMA as a distal tubular marker, and alkaline phosphatase with fast red as chromogen. BK viral DNA was detected in blood in 16 of 18 tested. Fourteen of 16 had cortex and medulla, and 2 had cortex only in the biopsy sample. Fourteen (100%) of 14 had double-positive EMA and TAg expression (EMA+TAg+) in medullary collecting ducts. T antigen was evident in loops of Henle in nephrectomies. Thirteen (81.3%) of 16 had EMA+TAg+, and 10 (62.5%) of 16 had CD10+TAg+ cortical tubular segments. CD10+TAg+ cells were observed in the parietal epithelium of Bowman's capsule in 3 biopsies (18.75%). T antigen was observed in 5.1% of CD10+ tubular profiles compared with 21% of EMA+ profiles in renal cortex (P < .0001). Polyoma virus TAg was observed in medullary collecting ducts, in distal and proximal cortical tubules, and in Bowman's capsule, in decreasing order of frequency. Loops of Henle also had TAg. This distribution suggests potential for an ascending route of infection of allograft tubules in PVN.