This study was conducted to assess the impact of dietary incorporation of Spirulina platensis and selenium nanoparticles (SeNPs) individually or in combinations on growth performance, antioxidant status, humoral immune response, and microbial populations in diet and ileum of heat-stressed broilers. Ross-308 one-day chicks (n = 450) were fed one of 9 experimental diets with five replicate cages in 2 phases for 35 d. The experimental diets were a control basal diet without supplementation or with 0.1 mg SeNPs, 0.2 mg SeNPs, 5 g Spirulina, 10 g Spirulina, 0.1 mg SeNPs + 5 g Spirulina, 0.1 mg SeNPs + 10 g Spirulina, 0.2 mg SeNPs + 5 g Spirulina and 0.2 mg SeNPs + 10 g Spirulina per kg diet. Dietary supplementation with Spirulina and SeNPs significantly (P < 0.05) increased body weight gain and European production efficiency factor. Serum GPx and SOD were significantly (P < 0.05) increased with dietary Spirulina and SeNPs supplementation, while, TBARS was decreased (P < 0.05). Circulating immunoglobulin IgM, IgA and IgG were increased in treated birds compared to the control ones, while the antibody titers to IBD, AIV, and NDV were not significantly altered. The results showed that SeNPs and Spirulina exhibited dose-dependent antimicrobial activities against ileal counts of total bacterial, total molds and yeast, coliform, E. coli, Salmonella spp. and Enterococcus spp. However, ileal populations of Lactic acid bacteria were increased with dietary Spirulina and SeNPs in a dose-dependent manner. The microbial load in broilers' diets was reduced by dietary incorporation of S. platensis and SeNPs. These results indicate that Spirulina and SeNPs can be potentially used as growth promoters and antioxidant, immunostimulant, and antimicrobial agents in heat-stressed broilers.