Alkaliphilic bacteria belonging to the genera Bacillus, Staphylococcus, Micrococcus, Pseudomonas and Arthrobacter isolated from sediment samples of the alkaline Lonar lake situated in the Buldhana District of Maharashtra State, India, were studied for the production of protease activity. Among the 54 isolates obtained, Arthrobacter ramosus and Bacillus alcalophilus exhibited high protease activity using soyacake as a sole source of carbon and nitrogen. Protease activity was optimum at 1% initial substrate concentration, at 30 °C and under shake culture condition for both organisms. The enzyme was thermostable (65 °C), stable at pH 12 and also active in the presence of commercial detergent. This enzyme removed blood stains from cotton fabric indicating its potential use in detergent formulations.