hERG channels mediating the rapid delayed rectifier K+ current (IKr) are important for normal ventricular repolarisation. In native cardiac tissues, the hERG 1a subunit co-assembles with a subunit encoded by an alternate transcript, “ERG1b” which has a shorter N-terminus and influences current kinetics (Jones et al, 2004). There is some evidence that hERG 1b may confer altered pharmacological properties on hERG 1a/1b compared to hERG 1a alone (Sale et al, 2008; Abi-Gerges et al, 2011). Some of this evidence derives from automated patch-clamp at room temperature (RT). As hERG kinetics show a complex temperature dependence (Vandenberg et al, 2006), we have sought to investigate comparative pharmacology of hERG 1a and hERG 1a/1b using manual patch clamp at physiological temperature and for some drugs at room temperature. HEK-293 cells stably expressing either hERG 1a alone or transiently co-transfected with hERG 1a and hERG 1b plasmid constructs were utilised. Effects of chloroquine and fluoxetine on hERG tail currents recorded at -40 mV (IhERG) carried either by hERG 1a or hERG 1a+1b were assessed using a standard voltage clamp protocol (every 12s a 2-s depolarising step to +20mV followed by a repolarising step to -40mV to elicit tail currents were applied). Most experiments were performed at 37oC; for fluoxetine (previously reported to be more potent against hERG 1a/1b than hERG 1a with automated patch clamp at room temperature (RT) (Abi-Gerges et al, 2011)), we compared RT and 37oC. For concentration-response relations for drug inhibition of IhERG tail at − 40 mV, four different concentrations of chloroquine and fluoxetine were tested (n=4 to 7 cells per concentration). We also examined the effect of a single concentration close to IC50 value for hERG 1a inhibiton by fluoxetine on IhERG1a and IhERG1a/1b elicited with a ventricular action potential voltage command at 37oC. At 37oC the sensitivity of IhERG tails to each of chloroquine and fluoxetine did not differ between IhERG1a and IhERG1a/1b. At 25oC the deactivation kinetics of both IhERG1a and IhERG1a/1b were slowed compared to 37oC. This was accompanied by a ~2.1 fold decrease in sensitivity to fluoxetine for IhERG1a/1b tails versus ~1.4 for IhERG1a tails at 25oC compared to 37oC. We also observed a marked temperature dependence of the time-course of IhERG1a inhibition by fluoxetine but this was not the case for IhERG1a/1b. Finally, we did not observe any difference in IhERG1a and IhERG1a /1b sensitivity to 1 μM fluoxetine at 37oC under AP clamp. Our data are consistent with modest differences between IhERG1a and IhERG1a/1b pharmacology. These differences vary between drugs and experimental temperatures.