The present study was designed to develop a technique for oocyte collection using an ultrasound-guided transvaginal approach in prepubertal calves too small to accommodate manual transrectal manipulation. A commercially available, 5 MHz, convex-array ultrasound transducer designed for intravaginal use in women was custom-modified for use in calves. In Experiment 1, calves 10 to 16 wk old (n = 10) were restrained in the standing position in an adjustable squeeze chute with regional anesthesia. In Experiment 2, the follicle aspiration procedure was performed in 6 wk-old calves (n = 20) in dorsal recumbency after tranquilization and caudal epidural anesthesia. Ovarian superstimulation was induced in half of the calves using 750 IU eCG (Experiment 1) or 200 mg Folltropin (Experiment 2). Consistent visualization of both ovaries using the transvaginal approach was accomplished after considerable practice. Two methods of ovarian immobilization were attempted, but both interfered with the ultrasound image and were consequently abandoned. The inability to immobilize the ovary resulted in attempts to spear the intended follicle "free-hand." The contents of follicles >or= 6 mm in diameter were aspirated, filtered, and oocytes were located using a stereomicroscope. Although ovarian superstimulation did not resolve the problem of ovary movement, the number of follicles of adequate size to aspirate was dramatically increased. Extremely sharp needles were found to be very important in the free-hand technique. A total of 232 follicles was aspirated, and 100 oocytes were collected (43%). In summary, a transvaginal ultrasound-guided technique for oocyte collection was developed for young calves in a standing position (10 to 16 wk of age) and dorsal recumbency (6 wk of age). Results demonstrate the potential utility of this approach for deriving oocytes from young calves.