Abstract Postoperative pain is a frequent complication associated with root canal treatment, especially during apical instrumentation of tooth with preexisting periradicular inflammation Objectives The aim of this clinical study was to evaluate the influence of the instrumentation techniques on the incidence and intensity of postoperative pain in single-visit root canal treatment. Material and Methods Ninety patients with single root/canal and non-vital pulps were included. The patients were assigned into 3 groups according to root canal instrumentation technique used; modified step-back, reciprocal, and rotational techniques. Root canal treatment was carried out in a single visit and the severity of postoperative pain was assessed via 4-point pain intensity scale. All the participants were called through the phone at 12, 24 and 48 h to obtain the pain scores. Data were analyzed through the Kruskal–Wallis test. Results There was significant difference between all groups (p<0.05). The modified step-back technique produced postoperative pain significantly lower than the rotational (p=0.018) and reciprocal (p=0.020) techniques. No difference was found between the reciprocal and rotational techniques (p=0.868). Postoperative pain in the first 12 h period (p=0.763) and in the 24 h period (p=0.147) was not significantly different between the groups. However, the difference in the 48 h period was statistically different between the groups (p=0.040). Conclusion All instrumentation techniques caused postoperative pain. The modified step-back technique produced less pain compared to the rotational and reciprocal techniques.