Thymidine kinase 1 (TK1) is a cell cycle regulated enzyme with maximum expression during the S phase. Serum TK1 (S-TK1) is a unique biomarker for cell proliferation. Here, an optimized [(3)H]-thymidine (dThd) phosphorylation assay is described, which is as sensitive as the commercially available TK-REA and TK-Liaison assays for measurement of S-TK1 activity in dogs and humans. Serum samples from dogs (35 healthy, 32 with lymphoma, 2 with leukemia, and 35 with solid tumors) and humans (18 healthy, 9 with chronic lymphocytic leukemia, 10 with myelodysplastic syndrome) were analyzed using the [(3)H]-dThd assay. Mean S-TK1 activities were 1.11 ± 0.46 pmol/min/mL in healthy dogs and 1.15 ± 0.32 pmol/min/mL in healthy humans. S-TK1 activities in dogs with hematological malignancies were 24.2 ± 47.9 pmol/min/mL, and the receiver operating characteristic curve showed an area under the curve of 0.88. With a cut-off value of 1.9 pmol/min/mL (mean value ± 2 SD), the sensitivity was 0.94 and the specificity was 0.68. Very similar results were obtained with human samples (healthy and lymphoma cases). S-TK1 activities measured during chemotherapy of six dogs with lymphoma were drastically reduced. In one case, S-TK1 activity increased prior to relapse. S-TK1 levels in dogs with solid tumors did not differ from the healthy group. S-TK1 activities correlated with those determined with the TK-REA and TK-Liaison assays (r=0.92 and r=0.96, respectively). In conclusion, this optimized [(3)H]-dThd assay is fast, sensitive and economical for measuring S-TK1 activity and should increase its clinical use as biomarker.
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