Different nutritional and cultural factors were studied to determine the optimum conditions for isoamylase production by Pseudomonas sp. in a batch culture of the production medium. These factors include carbon, nitrogen and phosphate sources and their concentrations, temperature and pH. Results showed that the optimum conditions for isoamylase production by Pseudomonas sp. were achieved when the production medium was supplemented with maltose 1%, peptone 0.4%, and K2HPO4 0.4% as a carbon, nitrogen, and phosphate sources respectively, at initial medium pH 6, and incubation at 28°C for 24 hours. Under these conditions isoamylase productivity reaches the maximum, at which enzyme specific activity was 0.85 U/mg proteins.