The essential challenges in practical applications of date palm micropropagation include explant browning, reduced callus growth, low multiplication rate, and frequent tissue contamination. Our study aimed to evaluate the effects of cefotaxime (Cefo) and silver nanoparticles (AgNPs) on microbial contamination removal, callus growth, and shoot regeneration in date palm micropropagation. Cultures were initiated from bud tips of ‘Barhee’ date palm. Murashige and Skoog medium with auxins, cytokinins, and activated charcoal was amended with Cefo (50, 100, and 200 mg·l−1), AgNPs (0.125 and 0.250 mg·l−1), and a combination of both. The medium supplemented with 200 mg·l−1 Cefo and 0.250 mg·l−1 AgNPs resulted in the best callus growth (318 mg). In contrast, 200 Cefo + 0.125 AgNPs resulted in maximum organogenesis and shoot number per jar (83.4% and 14.1 shoots per jar, respectively). These two combinations resulted in contaminant-free cultures. The total amount of phenolic compounds was significantly reduced to 0.79 and 0.57 mg GAE·g−1 DM in shoots cultured in the above media, which was reflected in the low browning rate. The data revealed that the maximum endogenous IAA content of shoots (2.681 μg·g−1 and 2.345 μg·g−1) was obtained in response to 200 mg·l−1 Cefo + 0.250 AgNPs and 200 mg·l−1 Cefo + 0.125 mg·l−1 AgNPs, respectively. Therefore, the optimized compositions established in the present study could be applicable in reducing contamination and helping callus production and multiple shoot regeneration. To our knowledge, this is the first study of the antibacterial and growth-promoting effects of Cefo in combination with AgNPs in in vitro cultures of ‘Barhee’ date palm.