Abstract: The detailed kinetic study of the immobilized enzyme is usually important and pertinent for the better management of the enzyme preparation. Therefore, the suitable comparison of the various kinetic parameters of the immobilized α-amylase (on gelatin) with that of the soluble enzyme has been conducted. The purified α-amylase was immobilized on gelatin using glutaraldehyde as an organic hardener with an optimum immobilization of 82.5% followed by characterization with respect to pH, temperature, substrate concentration, effect of thiols and thiol inhibitors. The optimum pH shifted from 5.5 to 7.0 i.e., towards the basic side upon immobilization. The optimum temperature of gelatin immobilized α-amylase was 85°C as compared to soluble enzyme (75°C) showing an improvement in thermal stability. A decrease in Michaelis constant, Km from 2.427 to 1.870 mg mL-1 upon immobilization was observed. An increase in activity with all the thiols was observed. No marked inhibition with the thiol inhibitors (similar to the soluble enzyme) is an indication of the fact that there are no free thiol or sulfhydryl groups in soybean α-amylase which are necessary for catalysis. The reutilization capacity of the immobilized enzyme was up to seven cycles. The potential utility of the gelatin immobilized enzyme in removal of starch stain from cloths by various commercially available detergents and sodium lauryl sulphate was tested. The immobilized α-amylase could be considered as a potential candidate for use as a cleaning additive in detergents in order to facilitate the removal of starch stains due to which it may find potential application in laundry detergents.