There are limited data on the use of steroids and antibiotics in assisted reproductive technology (ART). Our aim was to evaluate the impact of these treatments on the outcome of IVF cycles in which Assisted Hatching (AH) was performed. We studied a retrospective cohort in a large university-affiliated infertility centre. Data from 1126 AH cycles performed between 2007 and 2009 were reviewed. Cycles were categorized as "treatment" (n = 640) and "no treatment" (n = 486), depending on whether they received steroids and antibiotics. The primary outcome was live birth. Secondary outcomes included implantation, spontaneous abortion, biochemical, clinical and ectopic pregnancy. Logistic regression was used to calculate odds ratios (OR) and 95% confidence intervals (CI). OR were adjusted (AOR) for age, BMI, baseline FSH, peak estradiol, cycle number, number of oocytes retrieved, number of embryos that underwent AH, number of high-implantation potential embryos, number of embryos transferred and physician in charge. The AOR (95% CI) of live birth was 1.91 (1.08-3.38), of clinical pregnancy, 1.75 (1.08-2.83) and of biochemical pregnancy, 0.24 (0.07-0.85). Our study suggests that treatment with steroids and antibiotics during AH cycles significantly increases the odds of live birth.