Biomonitoring of DNA damage by alkaline filter elution

Abstract

The majority of DNA lesions resulting from interactions of carcinogens with DNA are usually either single strand breaks or lesions which are converted to single strand breaks by treatment of DNA with alkaline solutions. A sensitive method of detecting DNA single strand breaks is the alkaline filter elution of DNA. We started to test this method for biomonitoring occupational exposure with sensitive experimental conditions using pH 12.6, where most alkali-labile DNA lesions are converted to single strand breaks. Under our conditions statistically significant differences can be detected between the elution rates of untreated V79 cells and cells treated with [3H]-thymidine 24 h prior to the elution. Statistically significant increases were detected in the elution rates of male smoking automobile mechanics and male smoking painters compared to non-smoking controls. No statistically significant differences were detected in the elution rates of male non-smoking automobile mechanics and male workers with a suspected exposure to halogenated aromatics compared to male controls. No statistically significant differences were observed in the elution rates of female smoking dry-cleaning workers compared to female smoking controls. Our experience showed that the alkaline elution technique can be a valuable tool for monitoring DNA damage in peripheral lymphocytes in man.

Biomarkers of genetic damage and inflammation in blood and bronchoalveolar lavage fluid among former German uranium miners: a pilot study

“…Alkaline filter elution Alkaline filter elution was performed and interpreted as described previously [9,10]. Aliquots of the lymphocyte suspension were lysed in the presence of proteinase K (0.5 mg/ml) for 30 min at room temperature.…”

DNA single strand breakage, DNA adducts, and sister chromatid exchange in lymphocytes and phenanthrene and pyrene metabolites in urine of coke oven workers.

“…Alkaline filter elution was carried out with a slight modification of the method described by Doerjer et al 2 A suspension containing a million cells was poured on to each filter (polycarbonate filter; pore diameter: 2 0 tiM; Nucleopore, Pleasanton, CA, USA). Cells were lysed with 3 ml 2 M NaCl, 10 mM Na2-EDTA, 0 5% Triton X 100, pH 10 with or without 0 5 mg/ml proteinase K for 60 minutes and then washed with 10 mM Na2-EDTA, pH 10 These results were used to calculate the DNA retention (D) of the filter as a percentage of the total amount of DNA; the elution rate (k) was calculated from k = 2 log D. The relative elution rates (krei) were calculated from the elution rates obtained for the V79 (kv79) cells, which were measured simultaneously (krei = k / kV79).'…”

Detection of DNA single‐strand breaks and glutathione in mononuclear blood cells of radiotherapy technicians

“…11,[17][18][19] The alkaline elution assay has been proven to be very sensitive in the detection of DNA lesions resulting from exposure to various chemicals in vivo and in tissue culture. 20 Circulating lymphocytes offer a number of advantages for performing genotoxicity monitoring. They are easily obtainable and can be considered as a non-organ specific internal monitor because they circulate throughout the body.…”