Pyruvate assay kit

CD3⁺TIGIT⁺ T or CD3⁺TIGIT^- T cells (2*10⁵/well) were stimulated with aCD3/CD28 for 8 hours and then cultured with fresh complete medium containing glucose. Lactate concentrations and pyruvate concentrations were analyzed according to the manufacturer’s instructions of the Lactate Assay Kit (Abnova, USA) and Pyruvate Assay Kit (Abnova, USA).

Pyruvate assay kit (Abnova), succinate colorimetric assay kit (BioVision), and acetate colorimetric assay kit (Sigma) were used for metabolite quantification. Procedures were followed according to the manuals. M. bovis BCG was grown in 6% glycerol Sauton’s medium for 9 or 11 days before culture filtrates were harvested for extracellular metabolite quantification. In some experiments, cells were pelleted and lysed by sonication prior to intracellular metabolite quantification. The concentrations of these metabolites were calculated based on their corresponding standard curves and normalized by protein concentrations.
Glutamate/glutamine was measured using the glutamine/ glutamate-Glo^TM assay kit (Promega) according to the manual. The glutamate/glutamine level [measured as relative light unit (RLU)] was recorded by Lumat LB 9507. Data were normalized by OD600.