Western blot analysis of the ratio pAMPK/AMPK. (A) Left: Representative western blot of AMPK and pAMPK in HEK293T cells under control conditions and following 4-h incubation in AICAR-containing solution (1 mM). Right: Quantification of active AMPK (pAMPK) protein expression related to the total AMPK (unpaired t test, P = 0.038, n = 5). (B) Real-time PCR from HEK293T cells transfected with hHCN4 from three technical replicates, indicating no significant fractional reduction in hHCN4 mRNA levels after AICAR treatment (paired t test, P = 0.5238, n.s.). Source data are available for this figure: SourceData FS1.