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Comparison of the alkaline comet assay results on fresh and one year frozen whole blood

We have already demonstrated that alkaline comet assay results in blood samples frozen as smal-volume aliquots with no cryoprotectors for short period can be comparable with those infresh blood samples. This time we wanted to compare the results after one year of freezing (-80ºC). Tail length and tail intensity parameters were measured in fresh and thawed whole blood samples taken from 30 healthy volunteers (13 males and 17 females). Medical surveillance was done in Merkur University Hospital Zagreb, Croatia, and all volunteers agreed to fill the questionnaire about their life-style habits, health condition and afterwards pass the medical checkup. The medical checkup consisted of the doctors physical examination, talk and blood and urine sampling to measure general laboratory parameters and also to perform the assay on their lymphocytes for DNA stability. Furthermore, due to time consuming analysis of individual slides, we have also compared the results of thawed blood on individual slides with 6- well gel assay (lab-made). The results demonstrated insignificantly higher DNA damage in thawed samples for tail intensity parameter, while the same was not seen for tail length. The damage observed in frozen samples correlated with the DNA damage measured in the freshly prepared samples. However, 6-well gel slide assay results on frozen blood were well correlated with individual slides on frozen blood, but no correlation was seen with the amount of DNA damage in the freshly prepared blood samples. Results indicate that comet assay can be successfully performed also in the frozen whole blood samples, but further clarification for high throughput 6/12/96 gel comet assay are needed not for comparison with frozen samples but with comparison with freshly samples and determination of inter-individual differences that can influence the results of the assay in fresh whole blood samples and the ones frozen for longer period of time.

comet assay ; diabetes ; frozen blood

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