Aim: Knowledge of root canal anatomy is crucial for successful root canal therapy. An in vitro study was performed to evaluate the number of roots, root canals and their types, level of division and fusion, curvature, location of apical foramina, transverse anastomosis and frequency of apical deltas in maxillary second premolars and to compare these findings with their radiographs. Materials and methods: A total of 90 extracted maxillary second premolars were collected and radiographed in a mesiodistal as well as buccolingual direction. The teeth were then decalcified, cleared and injected with haematoxylin dye. All of the specimens were examined under a stereomicroscope. The examination of the root canal system was based on Vertucci's classification. Results: In total, 94.4% of the teeth had one root and 5.6% had two roots. In cleared specimens, the percentage of Type I canals was 24.4%, Type II 34.4%, Type IV 27.8%, Type V 6.6% and Type VI 6.7%. The radiographic interpretation of the buccolingual view was found to be significantly different from the actual morphology (phi = 0.6, P 0.001) and the mesiodistal view provided most of the information. Conclusions: A careful study of the radiographs is important. Cone angulation techniques should be used for determining root canal complexities, so as to treat a maximal number of canals. This paper was presented at the 23rd FODI (Federation of Operative Dentistry of India) and 16th IES (Indian Endodontic Society) National Conference held in Kochin, India on 15 November, 2009. Keywords: canal morphology, decalcification, maxillary, radiographs, second premolar