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Clinical Chemistry 50: 1597-1606, 2004. First published July 9, 2004; 10.1373/clinchem.2003.030379
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(Clinical Chemistry. 2004;50:1597-1606.)
© 2004 American Association for Clinical Chemistry, Inc.

Cancer Diagnostics

Sensitive Nonradiometric Method for Determining Thymidine Kinase 1 Activity

Anders Öhrvik1, Maria Lindh1, Roland Einarsson1, Jacques Grassi2 and Staffan Eriksson3,a

1 DiaSorin AB, SE-161 02 Bromma, Sweden.
2 CEA, Pharmacology and Immunology Unit, CEA/Saclay, Gif sur Yvette, France.
3 Department of Molecular Biosciences, Swedish University of Agricultural Sciences, BMC, Uppsala, Sweden.

aAddress correspondence to this author at: Department of Molecular Biosciences, Swedish University of Agricultural Sciences, BMC, SE-75123 Uppsala, Sweden. Fax 46-18 550762; e-mail Staffan.Eriksson{at}vmk.slu.se.

Background: Thymidine kinase 1 (TK1) is a cytoplasmic enzyme, produced only in the S-phase of proliferating cells, that has potential as a tumor marker. Specific determination of TK1 in serum is difficult, in part because of differences in the physical properties of serum TK1 compared with cytoplasmic TK1.

Methods: The first step in the new assay was phosphorylation of 3'-azido-2',3'-deoxythymidine (AZT) to AZT 5'-monophosphate (AZTMP) by TK1 present in patient material. The AZTMP formed was measured in a competitive immunoassay with specific anti-AZTMP antibodies and AZTMP-labeled peroxidase. Results were compared with those of a TK radioenzyme assay (REA) for 78 samples from patients suffering from hematologic diseases.

Results: The detection limit was 78 µIU/L, and within-run CVs <20% were seen for samples with TK1 down to 130 µIU/L. Cross-determination of the mitochondrial isoenzyme TK2 activity was <0.1%. Between-assay imprecision (CV) was 3.5–7.4%, and the within-assay imprecision was 4.1–9.1%. In studies of recovery and linearity on dilution, measured values ranged from 84% to 115% of expected at concentrations of 0.26–10.4 mIU/L. Results of the new assay (mIU/L) = 0.109 x TK REA (U/L) + 0.092. Heterophilic antibodies did not interfere in the assay. The upper 95th percentile, in 100 healthy individuals, was 0.94 mIU/L, and the median value was 0.43 mIU/L.

Conclusion: The TK1 enzyme-labeled immunoassay uses a stable substrate, is precise, appears to be accurate, and is resistant to interferences. It may provide a practical tool in the management of hematologic malignancies.




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