An experiment was conducted at the Poultry Research Station of Tamil Nadu Veterinary and Animal Sciences University, Chennai, India to study the effect of in ovo injection of glucose, glutamine and egg white protein (EWP) on the production performance and serum biochemistry of broilers. On the 18th day of incubation, of total 500 eggs, keeping 100 eggs as control (T1), the remaining 400 eggs were grouped in to four each containing 100 eggs. They were randomly selected for injecting with normal saline (T2) or normal saline containing 5 % glucose (T2) or 0.4 % glutamine (T3) or 12% egg white protein (T4) or 5 % glucose + glutamine 0.4 % + egg white protein 12% (T5). A pin head size hole was made just below the air cell and 0.5 ml of (in ovo injection) nutrient solution was injected into the amnion using a insulin syringe with 31G needle (0.25 mm x 8 mm).The hole was sealed and the period of incubation was completed. After hatching, 400 chicks (80 chicks selected from each in ovo fed group) were randomized in to 5 treatments of 4 replicates with 20 chicks each. Data on hatchability, hatchling weight, body weight, weight gain, feed consumption, FCR, liveability and liver glycogen, serum glucose, total protein, albumin and globulin were recorded and subjected to statistical analysis. Results revealed that in ovo injection of glucose or egg white protein resulted in significant (P<0.05) increases in the hatchability, chick weight, body weight, FCR, liveability and liver glycogen, serum total protein, albumin and globulin concentrations. However, the feed consumption was significantly (P<0.01) increased in Glucose injected groups, sham and control as compared to other groups. In ovo injection of glucose or EWP either alone or in combination improved the productive performance of the broilers.