The aim of this study was to investigate whether the use of operating microscope in combination with ultrasonics increased the rate of second mesiobuccal (MB2) canal detection in permanent maxillary first molar teeth. A hundred extracted human maxillary first molars were assessed. After location of the main canals, the MB2 canal was sought in all teeth first without microscopy, then with the aid of the operating microscope and finally with the combined use of the operating microscope and ultrasonics. With these techniques, the MB2 canal was detected in 62%, 67% and 74% of the teeth, respectively. The combination of the operating microscope and ultrasonics detected significantly more MB2 canals than when no microscopy was utilized (P < 0.05). Sectioning of the roots disclosed the presence of the MB2 canal in 82% of the teeth. Twenty-nine per cent of the teeth had a separate MB2 canal orifice and separate apical foramina. The results of this study suggested that the combined use of the operating microscope and ultrasonics increased the detection of MB2 canals in maxillary first permanent molars.