Use of a simple light absorbance assay to measure lymphocyte proliferation

The proliferative response of human lymphocytes to stimuli such as foreign histocompatibility antigens or mitogens is generally assessed by measuring the amount of tritiated thymidine which the cells incorporated in culture. In this paper, the possibility of assessing lymphocyte proliferation and viability by an empirical assay, using measurement of light absorbance on a ELISA reader in the yellow wave length (450 nm/air-550 nm/air), has been studied. The correlation of these measurements with a colormetric viability assay using MTS/PMS, with tritiated thymidine incorporation and with trypan blue exclusion viability counting, was determined. The results showed that the light absorbance assay correlated well with cell proliferation during 48-120 hours culture period and with cell viability after a 72 hour period. The MTS/PMS colormetric assay as well as trypan blue exclusion cell counting confirmed that the light absorbance assay was not merely caused by dead cells. This data confirm that the light absorbance assay is sufficiently sensitive to low levels of proliferation to allow detection of such responses at least as effectively as thymidine incorporation. The light absorbance assay procedure avoids the expense, time and hazards associated with scintillation counting, and is simple to perform without the necessity for reagents and preparative steps required by other assays.