MOLECULAR DOSIMETRY BY FLOW CYTOMETRIC DETECTION OF THYMINE DIMERS INMONONUCLEAR-CELLS FROM EXTRACORPORALLY UV-IRRADIATED BLOOD

UV-induced DNA damage in mononuclear leucocytes can be quantified by f low cytometry of fluorescence from a labelled monoclonal antibody that specifically binds to thymine dimers (T<->T): specific fluorescence i s already detectable after exposures of 1-2 J m(-2) of 254 nm radiatio n and shows a linear relationship with dose. The distribution of UV fl uences over an irradiated volume can thus be ascertained by measuring the UV-induced T<->T loads of the individual cells from that volume. A fter irradiation of mononuclear cells in a phosphate buffer solution i n a Petri dish, most cells showed a similar intensity of specific T<-> T fluorescence, forming a single sharp peak in the fluorescence histog ram. This signifies an even distribution of fluences over the cells. I t was noticed, however, that a variable minor fraction of mononuclear cells (usually less than 10%) could be resistant to immunostaining; th is fraction was rejected from the calculation of the specific fluoresc ence. The flow cytometric technique was also applied to blood cells ex posed in an ISOLDA device, which is in use in Russian clinics for UV i rradiation of whole blood for therapeutical purposes. Only a small fra ction of mononuclear cells in a sample of whole blood treated in ISOLD A acquired a detectable T<->T load after exposure to lamps which emit predominantly either WC or UVB light ((3.6+/-1.0)% and (1.8+/-0.4)% of all analysed cells respectively). This small fraction had received a large variation in fluences, resulting in differences in nuclear T<->T loads by a factor of 200. There had apparently not been enough mixing of cells during the UV irradiation of the blood to even out differenc es in UV fluences. Hence the trigger of the reported curative effects of reinfused, UV-irradiated blood would appear to originate from the i nitial photomodification of a minor fraction of the extracted blood vo lume (1-2 ml per kilogram of body mass).