Evaluation of X-radiation and UV-B radiation for elimination of [formula omitted]-thymidine incorporation into responder cells in the IL-2 producing helper T lymphocyte precursor assay

Abstract

The helper T lymphocyte precursor (HTLp) assay is of value for predicting graft-vs.-host disease after allogeneic bone marrow transplantation. The assay is based on limiting dilution analysis and requires detection of the amount of interleukin 2 (IL-2) produced by one activated T cell. IL-2 is detected by 3 H -thymidine ( 3 H -TdR) incorporation into the IL-2 dependent cell line, CTLL-2. Detection of IL-2 in the whole culture volume of the wells in the microtiter plates increases sensitivity, but requires elimination of 3 H -TdR incorporation into the responder cells in the HTLp assay. We have compared the ability of X-radiation and ultraviolet B (UV-B) radiation to eliminate 3 H -TdR incorporation. X-irradiation of the cells reduced 3 H -TdR incorporation by 90% with doses up to 400 Gy, but the incorporation was still 10 times higher than in wells with stimulator cells only. UV-B irradiation (with Philips TL 12 tubes) of the cells with ≥2 J/cm 2 decreased 3 H -TdR incorporation to the level of wells with stimulator cells only. Neither X-irradiation nor UV-B irradiation of the cultures affected IL-2 produced in the assay or human recombinant IL-2 measured by 3 H -TdR incorporation into the IL-2 dependent cell line, CTLL-2. HTLp frequencies determined after 25 Gy X-irradiation were higher (mean 1.5, range 1.02–2.2 times higher) than HTLp frequencies determined after UV-B irradiation with 2 J/cm 2.