HJC0350 is an effective and specific EPAC2 inhibitor (IC50: 0.3 μM), and no inhibition of Epac1.

EPAC2:0.3 μM

In Vitro IRE1 RNase and RIDD Assays: Analysis of radiolabeled Xbp1 substrate cleavage is performed as previously except that mammalian IRE1 reaction buffer is used. In vitro RIDD substrates are synthesized by in vitro transcription using the T7-MAXIscript Kit in the presence of 32P ATP or Cy5-UTP on templates isolated by RT-PCR from mouse Min6 cells (Ins2) or PCR from cloned XBP1 cDNA. The resulting products are gel purified to obtain full-length substrate. Reactions are then separated by 15% UREA-PAGE for analysis by phosphorimaging or by near-infrared imaging using the LI-COR Odyssey scanner.