Animal feed and ingredients have been shown to be potential vectors of pathogenic bacteria in to the human food chain. Introduction of contaminated materials can lead to facility contamination, which can be easily spread to non-contaminated materials, including finished feeds. Upon detection of contamination, facility decontamination is necessary to prevent potential cross-contamination to feedstuffs. The purpose of this experiment was to evaluate a standardized protocol to decontaminate an animal feed manufacturing facility using Enteroccoccus faecium (ATCC 31282) as an indicator. A pelleted swine diet inoculated with E. faecium was manufactured at the Kansas State University Feed Safety Research Center. Environmental samples including swabs, RODAC plates and air samples were collected and consisted of: 1) baseline before inoculation, 2) after inoculated feed production, 3) after production physical clean using pressurized air, 4) after chemical clean with a quaternary ammonium-glutaraldehyde blend, 5) after chemical clean with sodium hypochlorite, 6) after facility heat-up to 60°C for 24 h, 7) 48 h, and 8) 72 h. Air samples collected on the exterior of the facility confirmed pathogen containment as E. faecium concentrations were equal to or lower than baseline levels at each sample location. Decontamination step and its associated interactions were the only factors that affected E. faecium incidence (P < 0.0001 vs. P > 0.22). After production of the inoculated diet, 85.7% of environmental samples were positive for E. faecium. Interestingly, physical cleaning had no effect on contamination (P = 0.32). Chemical cleaning with a quaternary ammonium-glutaraldehyde blend and sodium hypochlorite each reduced (P < 0.0001) E. faecium contamination to 28.6% and 2.4% of tested surfaces. All samples were negative after 48 h of heating. In summary, both wet chemical cleaning and facility heating resulted in substantial E. faecium decontamination, but not physical cleaning. In addition, this experiment confirmed both successful containment and decontamination of biological pathogens in the tested pilot-scale feed mill.