ABSTRACT Response surface methodology was used to optimize lincomycin production by Streptomyces lincolnensis NRRL ISP-5355 in submerged fermentation. Screening of fermentation medium components to find their relative effect on lincomycin production was done using Plackett-Burman design. Malt extract, dextrin, soluble starch and (NH4)2SO4 were the most significant nutrient influenced on lincomycin production. Central composite design was applied to determine optimal concentrations of these factors and the effect of their mutual interactions. The interaction between soluble starch and (NH4)2SO4 was found to enhance the production, whereas malt extract and dextrin exhibited an influence independent from the other two factors. Using this statistical optimization method, maximum lincomycin concentration of 1345 μg/ml was obtained which represented a 40.5 % increase in titer than that acquired from the non-optimized medium. This statistically optimized medium was employed for lincomycin production through immobilization of Streptomyces lincolnensis by adsorption on synthetic cotton fibers. Immobilization technique improved the concentration to 1350 μg/ml higher than that produced from free cells cultures and could be maintained for longer than 17 days in a repeated batch system.