In-ovo injection with SeNPs or natural antioxidants during the incubation period could boost the growth performance of hatchlings. The objective of this study was to evaluate the effect of in ovo injection of SeNPs or clove oil on the growth performance and immunological and physiological responses of boiler chicks. A total of 240 fertile eggs used in the current experiment were obtained from avian broiler breeder flocks. The eggs were randomly divided into four groups, with 60 eggs per group. The first group consisted of intact, non-injected eggs (negative control, NC), and the second treatment injected into the yolk sac with 0.1 ml saline ( positive control, PC.). While the 3rd and 4th treatment were injected with 0.1 ml SeNPs and clove oil. Hatched chicks from each group were randomly assigned to three replicates and reared until 35 d of age The results showed highly significant differences among groups in terms of chicks' weight at day old, growth performance, lymphoid organs, serum total antioxidant, and immunoglobulin G (IgG). Additionally, SeNPs and clove oil had a positive effect on the total bacterial count and pathogenic bacteria compared with the control groups. However, the use of SeNPs or clove oils resulted in non-significant differences in serum total protein, albumin, and globulin levels compared to the control groups. Furthermore, it can be concluded that in ovo injection of SeNPs and clove oil during the incubation period has a positive effect on broiler chicken weight at hatching and growth performance, as well as on the immunological and antioxidant status of broiler chicks.