Because of numerous indigenous microorganisms on excised teats that interfere with enumeration of challenge cultures on nonselective media, a series of experiments used various physical and chemical treatments to reduce overall numbers. Physical treatments of teats included autoclaving at 121 degrees C for 15 min, boiling in distilled water for 10 min, and heating by microwaves for 2 min. Chemical treatments included exposing teats to various concentrations of sodium hypochlorite, hydrogen peroxide, iodophor disinfectant, and combinations of the three. All treatments were unsuitable. Selective and differential media were screened for their ability to restrict bacterial contamination from excised teats while maintaining good recovery of Streptococcus agalactiae. Edwards and NPC agars were superior for recovery of S. agalactiae. Four methods for recovery of S. agalactiae from excised teats were evaluated. Hand agitation of teats in neutralizer and stomaching teats in neutralizer were equally superior to swabbing teats and rinsing teats with neutralizer expelled from a syringe. Studies of elapsed time demonstrated that 1 min or less between challenge exposure and teat dip exposure was just as suitable and more expedient than 5 min.