MR1 presents a protein-containing antigen from the mycobacterial cell wall.

<p>(A) CFP or CW from the Mtb strain H37Rv were added (5 µg/ml) to DCs (25,000/well) for 1 h prior to the addition of one of 21 NC clones (5,000/well) followed by IFN-γ ELISPOT assay. (B) DCs (25,000) loaded with CW overnight were incubated with anti-MR1 blocking antibody (clone 26.5) or a mouse IgG2a isotype control (both at 5 µg/ml) for 1 h prior to the addition of T-cell clones (10,000/well). (C) dCW from Mtb was treated with proteases (subtilisin, trypsin, chymotrypsin, pronase, Glu-C) and added (5 µg/ml) to DCs (25,000/well) for 1 h before the addition of 21 NC clones (5,000/well) that were tested for their ability to produce IFN-γ in an ELISPOT assay. Reversed phase- high performance liquid chromatography (RP-HPLC) chromatogram analyses were used to confirm the inactivation of proteases. No responses were detected in the absence of DCs. (D) DCs were infected with <i>S. typhimurium</i>, <i>L. monocytogenes</i>, <i>and S. aureus</i> for 1 h with a calculated moi of 145, 6, and 15, respectively. DCs were washed, antibiotics added, and DCs (25,000) were incubated with three different Mtb-reactive MAIT-cell clones (10,000/well) that were tested for their ability to produce IFN-γ in an ELISPOT assay. Results shown are similar to a minimum of three independent experiments where <i>S. typhimurium</i>, <i>L. monocytogenes</i>, <i>and S. aureus</i> were tested at a variety of moi ranging from 5 to 150.