Laparoscopy is now a reliable method for staging gastrointestinal cancer, orienting the therapy, and avoiding unnecessary laparotomy. Natural orifice transluminal endoscopic surgery (NOTES) is an emerging concept with potential advantages for patient recovery. The first case of clinical diagnostic application of transvaginal NOTES for diagnostic cancer staging is presented. Informed consent and Institutional Commission approval were obtained for transvaginal clinical trials. On February 28, 2007, a patient with elective surgical indication for diagnostic cancer staging was submitted to transvaginal NOTES procedure, and intra- and postoperative parameters were documented. In a 50-year-old female patient presenting with ascitis, diffuse abdominal pain, and weight loss for 2 months, diagnosis of peritoneal carcinomatosis was suspected, which was also found when a CT scan was performed. Transvaginal NOTES was used for diagnostic staging of the patient, using a colonoscope introduced into the abdomen through a small incision in the vagina. Biopsies of liver, diaphragm, ovaries, and peritoneum were successfully performed. Operative time was 105 min, vaginal access and closure was obtained in 15 min. Abdominal inventory was reliable, and all 16 biopsies taken were positive for ovarian adenocarcinoma. The patient was dismissed 48 hours after the procedure without complications. Recent literature and experience of the study group suggest possibilities for preliminary clinical applications by transvaginal natural orifice surgery for diagnostic purposes.