Mouse anti tg2 antibody

DMF and H₂N-ACR-immobilized FG beads were prepared by Tamagawa Seiki (Tokyo, Japan). Recombinant human TG2 (100 ng) was incubated on a rotator at 4 °C overnight with 5 µL FG beads immobilized with 0, 0.1, 0.3, or 1 mM H₂N-ACR in the absence or presence of 50 nmol free ACR. Beads were extensively washed with Tris buffer (pH 7.8) and heated for 5 min at 97 °C. The boiled and eluted sample was loaded onto sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to a PVDF membrane (Bio-Rad Laboratories, Hercules, CA, USA). After blocking, the membrane was probed with mouse anti-TG2 antibody (1:3,000 dilution, MA5-12739, Thermo Fisher Scientific) overnight at 4 °C. The resultant blots were incubated with HRP-conjugated secondary antibodies (1:2000 dilution) and detected using an Amersham ECL Plus western blotting Detection System (GE Healthcare, Piscataway, NJ, USA). Band intensities were quantified using the ImageJ software (National Institutes of Health, USA).