Objectives: Since it has been suggested that intracytoplasmic sperm injection (ICSI) might impair the early development of embryos and twin pregnancies have physiologically different aspects from singleton pregnancies, we aimed to investigate the effect of ICSI on the human chrionic gonadotropin (hCG) levels in twin pregnancies. Methods: A total of 220 clinical pregnancies which underwent controlled ovarian hyperstimulation and in vitro fertilization/ICSI were analyzed retrospectively. Results: In twin pregnancies, the mean number of retrieved oocytes and estradiol (E(2)) levels on hCG day were significantly higher in ICSI group in spite of lower dose of gonadotropin. The hCG levels according to insemination method did not demonstrate any difference after adjustment for age of patient and partner, BMI, treatment regimens, serum E(2) on hCG day, number of oocytes retrieved and date of sampling. Conclusions: The hCG values according to fertilization methods were not significantly different both in twin pregnancies with adjustment for confounders.