Gamete collection, gamete preservation, hormone analysis, and artificial insemination have become integral parts of in situ and ex situ conservation programs for threatened and endangered species. Although these methods have been used to assist conservation in many different vertebrate groups, limited work has been done in reptiles. For example, semen collection in lizards has only recently been described and determined to be safe. The purpose of this study was to determine effective doses of human chorionic gonadotropin (hCG) for increasing plasma testosterone concentrations in the veiled chameleon (Chamaeleo calyptratus), as well as to determine whether the rise in testosterone impacts semen collection. A crossover design was used. Initially, chameleons were given injections of 100, 200, and 300 IU of hCG per animal, and serial plasma testosterone measurements were collected over 24 h. The 100-IU/animal dose was determined to increase plasma testosterone concentrations at a similar level to that of the 200- and 300-IU/animal doses. Next, we determined weekly injections of hCG (100 IU/animal) would maintain elevated testosterone concentrations over 30 days. In addition, we determined that elevated testosterone secondary to repeated injections of hCG decreased testicular size as determined by ultrasound. Repeated hCG injections and long-term elevation of plasma testosterone concentrations did not increase the likelihood to collect a semen sample with electroejaculation or improve ejaculate quality. Further research is needed to exogenously stimulate spermatogenesis and increase ejaculate quality to perform timed semen collections in male reptile species.