Figure 3. Analysis of NAD⁺ metabolites in the supernatant of Jurkat/CD73⁻ cells. (A and B) The enzymatic conversion of NAD⁺ to adenosine diphosphate ribose (ADPR) (a CD38-dependent reaction) and to AMP (CD203a-dependent) was monitored by high-pressure liquid chromatography (HPLC). Representative resting (A) and phorbol 12-myristate 13-acetate (PMA)-activated-activated (B) Jurkat/CD73⁻ cells upon incubation for 60 min at 37°C with 100 µM NAD⁺ are shown. The identity of peaks was confirmed by the co-migration and absorbance spectra of reference standards using a retention time (R_t) window of ± 5%. A representative plot of retention time of NAD⁺ metabolites in a single HPLC run is depicted on the right.