This study evaluated the impact of in ovo injection of L-Arginine (L-Arg) on the hatchability, physiological and post-hatching parameters Cobb 500 broiler chicks. Three hundred and eighty fertilized eggs from Cobb 500 strain were randomly distributed into four-treatment groups. The 1^st group (110 eggs, non-injected negative control, T₁), the 2^nd group (110 eggs, injected with distilled water positive control, T₂), the 3^rd group (80 eggs, injected with 1% L-Arg, T₃), the 4^th group (80 eggs, injected with 3% L-Arg, T₄). The embryonic amnion in the T₃ and T₄ groups was injected with 0.5 mL of L-Arg solution on day 18.5 of incubation. There were significant differences (P0.01) in hatchability and growth performance between the treatment-groups. T₃ had a higher percentage of hatchability (97.48%) than T₂ (94.57%). T₃ also had the lowest late death rate (1.23%). T₄ chicks were superior (P≤0.01) in live body weights, weight gains and feed consumption at different ages than those of T₃ and control chicks. Chicks hatched from eggs injected with 3% L-Arginine had higher (P≤0.01) overall means of the proportional weight of dressing, breast, heart, hips, gizzard, caeca, ileum, large intestine, duodenum length, ileum length, caeca length, and dressing percentage than chicks produced from the control groups. On day 14, the proportional weights of the spleen, bursa of Fabricius, and thymus were significantly higher in the ovo injection groups treated with 1% and 3% L-Arginine than in the control groups (P≤0.01). In comparison to other groups, the chicks hatched from eggs injected with L-Arginine in the T₃ and T₄ groups exhibited significant increases (P0.01) in serum IgG concentration. However, there were significant decreases (P0.01) in serum SGOT, SGPT, total protein, and globulin. T₄ chicks had higher values of Hb, PCV, MCV, MCHC and WBCs than the other groups at the 7^th day of age. Conclusively, the in ovo injection of L-Arginine treatments resulted in improvements to the hatchability, growth rate, blood parameters, and IgG concentration of broiler chicks