With the goal of optimizing protocols, fixed-time artificial insemination (FTAI) was performed under field conditions in a study to evaluate conception rates in Girolando (5/8) heifers (18 to 24 months of age) treated with estradiol benzoate (BE) to induce ovulation. Forty heifers were allocated to 2 groups with 20 animals each: GI-sexed semen (2 million spermatozoa/dose) and GII-nonsexed semen (20 million spermatozoa/dose), with body condition score (BCS) of 3.0 ± 0.14 (scale of 1 to 5), middleweight of 335 ± 38 kg, so that it was made for reutilization of progesterone device. On Day 0 (morning), the animals received 2.0 mg of BE (RIC-BE^®,Tecnopec, São Paulo, Brazil), 0.075 mg of D-cloprostenol (Prolise^®, ARSA S.R.L., Buenos Aires, Argentina), and an intravaginal device of 1 g of progesterone (Primer^®, Tecnopec). On Day 8 (morning) the device was removed and 0.075 mg of D-cloprostenol was managed in the GI animals, then these animals received 1.0 mg of BE 24 h after device removal. The insemination was in the late afternoon of Day 10 with a 6-hour delay (4 to 6 pm). The GII animals received Primer previously used for 8 days (being used in the GI animals under the same protocol). After pregnancy diagnosis at 60 days, a third group was created (GIII) using the not-pregnant animals of GI and GII. In GIII, Primer from third use [disinfected in solution Kilol^®-L 1:250 (Quinabra, São Paulo, Brazil) and rinsed with water, dried, and packed in plastic sacks and kept for 60 days] was used following the GI protocol with sexed semen. There were no significant differences (P > 0.05) in the conception rates of 40, 58, and 65% among groups GI (8/20), GII (11/20), and GIII (13/20). These results indicate very good conception rates for heifers, contrary to recommendations of commercial companies of sexed semen that alert for the conception rate for mated animals to be lower when using FTAI and mating protocols in mass, compared with that of AI after estrus observation.