Purpose: To determine the diagnostic utility of a frozen section biopsy in patients undergoing endoscopic submucosal dissection (ESD) for early gastric neoplasms with obscure margins even with chromoendoscopy using acetic acid and indigo carmine (AI chromoendoscopy). Materials and Methods: The lateral spread of early gastric neoplasms was unclear even following AI chromoendoscopy in 38 patients who underwent ESD between June 2007 and May 2011. Frozen section biopsies were obtained by agreement of the degree of lateral spread between two endoscopists. Thus, frozen section biopsies were obtained from 23 patients (FBx group) and not in the other 15 patients (AI group). Results: No significant differences were observed for size, histology, invasive depth, and location of lesions between the AI and FBx groups. No false positive or false negative results were observed in the frozen section diagnoses. Adenocarcinoma was revealed in three patients and tubular adenoma in one, thereby changing the delineation of lesion extent and achieving free lateral margins. The rates of free lateral resection margins and curative resection were significantly higher in the FBx group than those in the AI group. Conclusions: Frozen section biopsy can help endoscopists perform more safe and accurate ESD in patients with early gastric neoplasm.