Background: Solus One Listeria is designed to accurately detect Listeria species (Listeria grayi, L. innocua, L. ivanovii, L. marthii, L. monocytogenes, L. seeligeri, and L. welshimeri) from stainless steel and plastic environmental surface matrixes using an antibody-based technology test system paired with proprietary SOLO+ media and combined with manual or automated sample preparation method. Objective: Solus One Listeria was evaluated for inclusivity and exclusivity and a matrix comparison study for two environmental surfaces. Methods: Solus One Listeria was compared with the following reference method for the method comparison study: the U.S. Food and Drug Administration Bacteriological Analytical Manual Chapter 10 from stainless steel and plastic environmental surfaces. Both the manual and automated preparation methods were performed for stainless steel and plastic environmental surfaces. Results: For the inclusivity and exclusivity evaluation, Solus One Listeria correctly identified all 50 target organism isolates and correctly excluded all 30 nontarget strains that were analyzed. In the method comparison study, both Solus One Listeria manual and automated preparation methods demonstrated no significant differences based on probability of detection statistical analysis between presumptive and confirmed results or between candidate and reference method results for two environmental surfaces after 22-30 h of enrichment time. Probability of detection analysis of Solus One Listeria method robustness, product consistency (lot-to-lot), and stability studies using the automated preparation method demonstrated no statistically significant differences. Conclusions: The data from the study support the product claims of Solus One Listeria for the accurate detection of Listeria species, using both the manual and automated methods (using the Dynex DS2 instrument), on both environmental surfaces analyzed.