Maintaining similar memories in a distinct and nonoverlapping manner, known as pattern separation, is an important mnemonic process. The medial temporal lobe, especially the hippocampus, has been implicated in this crucial memory function. The present study thus examines whether it is possible to modulate pattern separation using bilateral transcranial direct current stimulation (tDCS) over the temporal lobes. Specifically, in this study, pattern separation was assessed using the Mnemonic Similarity Task following 15-min offline bilateral temporal lobe tDCS (left cathode and right anode or left anode and right cathode) or sham stimulation. In the Mnemonic Similarity Task, participants studied a series of sequentially presented visual objects. In the subsequent recognition memory test, participants viewed a series of sequentially presented objects that could be old images from study, novel foils, or lures that were visually similar to the studied images. Participants reported whether these images were exactly the same as, similar to, or different from the studied images. Following both active tDCS conditions, participants were less likely to identify lures as ‘similar’ compared with the sham condition, indicating a reduction in pattern separation resulting from temporal lobe tDCS. In contrast, no significant difference in overall accuracy was found for participants’ discrimination of old and new images. Together, these results suggest that temporal lobe tDCS can selectively modulate the pattern separation function without changing participants’ baseline recognition memory performance.