Seven isonitrogenous (400 g/kg crude protein) and isocaloric (17.89 kJ/g gross energy) purified diets (casein-gelatin based) with different concentrations of calcium (0, 1, 2, 3, 4, 5 and 6 g/kg) supplemented with calcium lactate at the levels of 0, 5.4, 10.8, 16.3, 21.8, 27.27 and 32.73 g/kg were prepared and fed for 12 weeks to triplicate groups of Heteropneustes fossilis (7.46 ± 0.03 g) to determine the optimum dietary calcium requirement. Analysed values of calcium in the diets were 2.41, 3.82, 4.56, 5.99, 6.71, 7.40 and 8.19 g/kg, respectively. Absolute weight gain, specific growth rate, protein retention efficiency, protein gain and feed conversion ratio of fish fed diets with increasing levels of dietary calcium improved up to 5.99 g/kg and then levelled off. Whole-body protein, moisture and ash contents improved up to 5.99 g/kg dietary calcium and stabilized thereafter. However, whole-body fat exhibited reverse pattern and decreased with incremental levels of dietary calcium up to 5.99 g/kg. Whole-body and vertebrae mineralization was also significantly affected (p < .05) by the increasing dietary calcium levels. Alkaline phosphatase activity improved significantly (p < .05) up to 6.71 g/kg, and no change was recorded beyond this level. Serum calcium and phosphorus concentrations were not influenced (p > .05) by dietary calcium levels. The Ca-P ratio remained static in the whole body, vertebrae and serum. Broken-line regression analysis of data obtained on growth, mineralization and serum ALP activity against increasing levels of dietary calcium reflected the optimum calcium requirement between 5.77-6.81 g/kg diet.