Estrus detection and determination of time of insemination are very important factors in reproduction management of cattle. Therefore an estrus synchronization schedule in combination with induction of ovulation and a single insemination at a predetermined time in dairy cattle was established to achieve high pregnancy rates (Kanitz et al. 2002 Reprod. Nutr. Dev. 42, 587–599; Becker et al. 2004). The aim of the recent study was to investigate the influence of the number of spermatozoa per insemination dosage on embryo development and the interrelationship between number of accessory sperms per embryo and its development using this schedule. In total 116 German Holstein heifers received GnRH (i.m.; 0.05 mg Gonavet^®, Veyx, Schwarzenborn, Germany) 60 h after PGF2α application (i.m.; 0.5 mg Cloprostenol forte^®, Jenapharm, Jena, Germany) administered between Days 8 and 14 of the estrous cycle. Artificial insemination was carried out 13 h after GnRH application. Three different dosages of spermatozoa (15 × 10⁶, 5 × 10⁶, and 1 × 10⁶) from three ejaculates from four fertile bulls were used. Embryos and oocytes were flushed from the oviducts of animals ovulated (n = 106; ovulation rate 91.3%). Animals were slaughtered on Day 4 after insemination. The quality of the embryos and oocytes was evaluated by microscopic examination. Embryos were stained with Hoechst 33258 to verify the number of accessory sperm. The evaluation of the data was carried out with the GLM procedure of the statistics software package SAS^® (SAS Institute, Inc., Cary, NC, USA). As a post-hoc test the Student’s t-test was used. Significance was set at P = 0.05. After flushing of 106 animals, 85 embryos and oocytes was recovered (recovery rate 80.2%). Relative to the three sperm concentrations, there were no significant differences among fertilization rates (92.3, 96.2, and 78.8%) and among the portions of normal developed embryos (84.6, 80.7, and 75.8%, all respectively) between groups. Interestingly, significant differences were found according to the mean number of accessory sperm/embryo (29.6 ± 8.4, 45.3 ± 8.6, and 6.5 ± 7.2, respectively) and in the portion of embryos without or with more than 10 accessory sperm/embryo. Results show that fixed-time insemination, independent of detection of onset of estrus can result in high fertilization rates. Insemination with dosages <5 × 10⁶ spermatozoa can reduce fertilization rates. Likewise, significant differences regarding fertilization rate were found after insemination of reduced sperm dosages of individual bulls. The number of accessory sperms/embryo seems to be an irrelevant parameter for quality of embryos produced under described conditions.