Nalidixic acid (nal) susceptibility of clinical isolates of Serratia marcescens from urinary tract infections, wounds, and blood was determined by the standard disk method, and results were compared with results obtained by a standardized tube dilution procedure using five different broth media. An attempt was made to approximate in vivo conditions by using urine as a test medium and correlating the activity of nal in urine with its activity in nutrient, Trypticase soy, brain-heart infusion, and Mueller-Hinton broths. The isolates were consistently susceptible to lower concentrations of nal in nutrient broth than in the other media. The results showed that Mueller-Hinton broth most closely reflected nal activity in urine. A standardized procedure for nal susceptibility testing of urinary tract isolates of S. marcescens is proposed.