Carcinoma of the endometrium is the most frequently diagnosed gynecologic malignancy in the western world. Because endometrial carcinoma is monoclonal in origin, the small samples obtained in endometrial pipelle biopsies can be used in PCR clonal studies to distinguish cancerous from noncancerous lesions. The method used for clonal analysis was based on RFLP of the X chromosome-linked phosphoglycerokinase gene and random inactivation of one X chromosome by methylation in women. Among 50 endometrial pipelle biopsies, 26 (52%) were found to be heterozygous for the above-mentioned polymorphism. Of the samples taken from these informative (ie, heterozygous) patients, six were monoclonal including five cases of endometrial carcinoma and one of endometrial atypical hyperplasia. In each case, the same pattern of monoclonality was present in the surgical specimen counterpart. All of the remaining samples were polyclonal and, when the anatomical pathology data were contrasted, they correlated with nonmalignant endometrium (five secretory, five proliferative, seven atrophic, and three simple hyperplasias). In addition, genetic alterations study of monoclonal endometrial samples revealed a K-ras point mutation and a c-erbB2/neu gene amplification in two different endometrial carcinomas. Both alterations were also detected in the surgical specimens. In addition, a diagnosed set of 10 samples of simple hyperplasia and 5 of atypical hyperplasia were subjected to clonal assay. Among eight informative cases, the three that showed that showed the monoclonal pattern corresponded with cases of atypical hyperplasia. No other genetic alterations were detected in these samples. In conclusion, our data indicate that the detection of clonality in endometrial biopsy samples obtained by pipelle would be a useful application for the early diagnosis of endometrial cancer.