Optimization of the T-cell proliferation assay in fascioliasis using a non-radioactive method, the Alamar Blue Assay

Abstract

T-cell proliferation studies are traditionally carried out with radioactive reagents or fluorescent reagents that require measurement with advanced technology instrumentation. We attempted to calibrate the optimal conditions suitable for the use of a non-radioactive assay for the measurement of a T-cell proliferation assay in bovine fascioliasis, but applicable to the study of other infectious diseases in our developing 'country setting, Crude antigen extract was prepared from 15 adult Fasciola gigantica flukes: Cellular responses were detected by the proliferatjon of peripheral blood mononuclear cells (PBMC) in response to stimulation by serial dilutions of the crude antigen extract. The results showed that the antigen dilution 1:1,600 gave the highest PBMC proliferative response (Stimulation Index, S.I = 1.10± 0:2). Percentage reduced Alamar Blue was 27.3-71.6%. This suggests that the cell-mediated immune response in bovine immunity to Fasciola infection may be reliably measured in our setting with the Alamar Blue Assay.