Comparison of whole blood and PBMC assays for T-cell functional analysis

Tuberculosis remains the foremost cause of morbidity and mortality, more than any other single infectious disease in the world. Cell mediated immune response plays a crucial role in the control of tuberculosis. Therefore, measuring cell mediated immune response against the antigens is having a vital role in understanding the pathogenesis of tuberculosis, which will also help in the diagnosis of and vaccination for tuberculosis.

The aim of the present study was to compare and optimize the assay conditions to measure the cell mediated immune response against M. tuberculosis specific antigens. Because the conventional PBMC assays (due to requirement of large volume of blood sample) are unable to screen more number of antigens within the same blood sample. So, here we have compared 6 days culture supernatants of 1:5 and 1:10 diluted blood and PBMCs from healthy laboratory volunteers, to assess the proliferative response of T lymphocytes and secreted IFN-γ levels against purified recombinant antigen of M. tuberculosis (MPT51, Rv3803c), crude antigens of M. tuberculosis (PPD) and mitogen (PHA).

We have observed good correlation between each assay and also the mean difference of these assays did not reach the statistical significance (p>0.05). From these results, we conclude that 1:10 diluted whole-blood cultures can be well-suited as an alternative assay to measure cytokine production and lymphocyte proliferation in comparison to the conventional PBMC assays. Moreover, 1:10 diluted blood assays require less volume of blood when compared to PBMC assays which will be useful particularly in paediatric and field studies in endemic countries, where blood volume is a limiting factor.