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Background. There are few studies of flow cytometric analysis for DNA heterogeneity of patients with superficial carcinoma of the esophagus limited to the epithelium or superficially invading the lamina propria or submucosa. Methods. Flow cytometric analysis of cellular DNA content was performed on superficial carcinomas of the esophagus using paraffin embedded blocks of the surgically resected specimens from 56 patients. To evaluate the intratumoral DNA heterogeneity, a total of 141 samples of the 56 tumors were analyzed, depending upon the tumor size. Results. One or two of the samples was available from 18 of 19 patients with tumors 2 cm or less in greatest dimension, whereas more than three of the samples were available from 22 of 37 patients with tumors 2.1 cm or greater in dimension (P < 0.003). Of 56 tumors, 40 (71.4%) exhibited DNA aneuploidy ; DNA heterogeneity was found in 26 tumors (46.4%). The remaining 16 tumors exhibited DNA diploidy. Two of the five tumors that were limited to the epithelium had DNA heterogeneity. The mean dimension of the tumors with DNA heterogeneity was significantly greater (5.8 ± 2.8 cm) than those exhibiting DNA diploidy (2.3 ± 1.1 cm) and DNA aneuploidy without heterogeneity (2.9 ± 2.4 cm). Recurrences after esophagectomy were detected in 6 of the 56 patients ; the DNA ploidy pattern of these six patients exhibited DNA heterogeneity. Conclusion. The incidence of DNA heterogeneity increases as tumor size increases and is associated with an increased risk of tumor recurrence after esophagectomy in patients with superficial carcinoma of the esophagus. Cancer 1995 ; 75 :914-9.