A biothermal activity detection method has been established to determine the potency of colistin.  The biothermal activity fingerprints of E. coli with colistin were determined.  There was a good linear relationship (r = 0.993) between logarithm concentration of colistin (lgC) and lag rate of growing time (Δ t %) when the  concentrations of colistin ranged from 17.0 to 41.6 u·mL⁻¹.  The average recovery rate was 100.3% (n = 9).  Using this method, there was no significant difference between results of colistin potency measurement and those using cup-plate method (P > 0.05).  As a result, biothermal activity detection method is sensitive, accurate, rapid, convenient and feasible to determine the potency of colistin.  This method can also be applied in real time and online to monitor the process of bacterial growth and could be complementary to the cup-plate method.