Background and Aim: Endoscopic definitions of Barrett's esophagus (BE) vary among countries, mainly because of the difficulty in diagnosing short-segment BE (SSBE) endoscopically. The aim of this study was to investigate whether the endoscopic identification of squamous islands and the specific position of columnar epithelium helps improve the diagnosis of SSBE. Methods: First, we prospectively enrolled 100 consecutive patients with SSBE and evaluated the number of identified squamous islands in the columnar epithelium with different modalities: white light (WL), narrow band imaging (NBI), and iodine chromoendoscopy. Second, in another group of 100 consecutive patients with tongue-like SSBE, the correlation of the location of Barrett's mucosa to the esophageal longitudinal folds (ridge or valley) was evaluated endoscopically. Results: It was possible to detect squamous islands in 48, 71, and 75 patients by WL, NBI, and iodine chromoendoscopy, respectively. The detection rate of squamous islands by NBI or iodine chromoendoscopy was significantly superior to that by WL. Tongue-like SSBEs were predominantly found on the ridge of mucosal folds (71%), similar to the location of mucosal breaks (84%). Conclusions: Squamous islands in the columnar epithelium were efficiently observed by NBI or iodine chromoendoscopy. SSBE was found more frequently on the ridges but not in the valleys of esophageal longitudinal mucosal folds. NBI endoscopic observation focusing on columnar epithelium with squamous islands on the ridges of distal esophageal folds may improve endoscopic detection of SSBE.