Different modified atmosphere conditions with various CO₂ concentrations on microbial load (psychrophile, mesophile aerobic and anaerobic bacteria) and physico-chemical properties of barramundi (Lates  calcarifer Bloch) fillets stored at 8°C were compared to determine the best packaging conditions. The gas conditions evaluated were 100% CO₂ (M₁), 75% CO₂/25% N₂ (M₂), 50% CO₂/50% N₂ (M₃), 25% CO₂/75% N₂ (M₄) and 100% N₂ (control). High CO₂ concentration (M₁ and M₂) of fish fillets delayed the psychrophilic bacteria growth compared to low CO₂ concentration (50% and 25% CO₂) as well as the mesophilic, anaerobic and histamine forming bacteria. We concluded that the atmosphere with 75:25% and 100:0 (CO₂:N₂) had the most appropriate gas composition to inhibit the microbial growth and prolong the shelf life of barramundi fillets (p<0.05).