Thymidine kinase assay

TK activity is assayed by a modification of the method of Breslow and Goldsby (1969) and has been described in detail (Clive et al., 19726). 

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An in vitro test with cytogenetic evaluation of chromosomal damage with mammalian cells OR an in vitro mouse lymphoma thymidine kinase assay... 

Mutagenesis Entacapone was mutagenic and clastogenic in the in vitro mouse lymphoma/thymidine kinase assay in the presence and absence of metabolic activation, and was clastogenic in cultured human lymphocytes in the presence of metabolic activation. 

Myhr B, Bowers L, Caspary WJ. 1985. Assays for the induction of gene mutations at the thymidine kinase locus in L5178Y mouse lymphoma cells in culture. In Ashby J, de Serres FJ, et al., eds. Progress in mutation research. Vol. 5. Evaluation of short-term tests for carcinogens. Amsterdam, The Netherlands Elsevier Science Publishers, 555-568. 

Clive, D., Flamm, W.G. and Patterson, J.B. (1972). A mutational assay system using the thymidine kinase locus in mouse lymphoma cells. Mutation Res. 16 77-87. 

Under Guideline S2B, the following standard test battery is recommended (1) a test for gene mutation in bacteria, (2) an in vitro test with cytogenetic evaluation of chromosomal damage with mammalian cells or an in vitro mouse lymphoma thymidine kinase (TK) assay, and (3) an in vivo test for chromosomal damage using rodent hematopoietic cells. 

The mouse lymphoma assay is in fact just one of several mammalian cell assays designed to determine increases in mutation rate. It focuses on the thymidine kinase (tk) assay in murine lymphoma cells (L5178Y), though tk data have also been produced from the human lymphoblastoid cell line TK6, and at the hgprt (hypox-anthine-guanine phosphoribosyl-transferase) locus in Chinese Hamster ovary or lung (V79) cells and mouse lymphoma cells. Since Ames mutation data and often... 

Styles, J.A., Clay, P. Cross, M.F. (1985) Assays for the induction of gene mutations at the thymidine kinase and the Na+/K+ ATPase loci in two different mouse lymphoma cell lines in culture. Prog. Mutat. Res., 5, 587-596... 

Wangenheim, J. Bolcsfoldi, G. (1988) Mouse lymphoma L5178 Y thymidine kinase locus assay of 50 compounds. Mutagenesis, 3, 193-205... 

The MLA is used for the detection of point mutations, structural aberrations and aneugenicity. The principle of the assay is that cells deficient in thymidine kinase (TK) due to the tk+/" or tk/ mutation are resistant to the cytotoxic effects of the pyrimidine analogue trifluoro-thymidine (TFT). TK proficient cells are sensitive to TFT. This influences the cellular metabolism and leads finally to an inhibition of further cell division. Thus mutant cells are able to proliferate in the presence of TFT, whereas normal cells, which contain TK, are not. The major advantage of the assay is its ability to detect a broad range of mutagenic events represented by optimal detection of both large and small colonies. 

The mouse lymphoma (MOLY) assay is an in vitro mammalian cell gene mutation test that can be used to detect gene mutations induced by chemical substances. The cell line used is the L5178Y MOLY cell. In these cell lines the most commonly used genetic end points measure mutation at the thymidine kinase (TK) locus on the mouse chromosome 11b. 

TK is an enzyme that allows cells to take up thymidine from the surrounding medium for incorporation into the DNA. Specifically for this assay, the TK phenotype is used. If a thymidine analog were added to suspension or soft agar, it would eventually be incorporated into the DNA, thereby resulting in the death of the cell. Cells deficient in thymidine kinase (TK ) due to the mutation (Tk+ are resistant to the cytotoxic ef-... 

Moore MM, Honma M, Clements J, et al. (2002) Mouse lymphoma thymidine kinase gene mutation assay followup International Workshop on Genotoxicity Test Procedures, New Orleans, Louisiana, April 2000. Environmental and Molecular Mutagenesis 40(4) 292-299. 

M. oleifera extracts inhibits plaque formation of anti-herpes simplex vims type 1 (HSV-1) more than 50% at 100 ag/ml in a plaque reduction assay (55). M. oleifera extracts are also effective against thymidine kinase-deficient HSV-1 and phosphonoacetate-resistant HSV-1 vims strains. The extract ofM. oleifera at a dose of 750 mg/kg body weight per day significantly delays the development of skin lesions, prolongs the mean survival times and reduces the mortality of HSV-1 infected mice. Compared to the synthetic compound acyclovir, M. oleifera extracts delay the development of skin lesions and has mean survival times as acyclovir. A polysaccharide from hot aqueous extract of mature pods (fruits) of M oleifera with a structural repeating unit [->4)-a-D-GlCp(l->] has immunoenhancing properties (76). 

Preparation of Crude Extracts The parasite larvae or adult forms were homogenized by grinding with sand in a mortar (placed on ice) with three to five volumes of ice-cold 0.05 M phosphate buffer, pH 7.5, containing 0.1 M KCl and 0.01 M 2-mercaptoethanol, except for assays of thymidine kinase activity, when phosphate buffer was substituted by Tris-HCl buffer at the same concentration and pH. The suspension was sonicated and centrifuged to obtain crude extract. 

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