The inclusion of rumen buffers in ruminant feeds has gained widespread adoption for the prevention of rumen acidosis, thereby avoiding the negative production and health consequences of low rumen pH and resulting in improved feed efficiency. Benchmarking and quality controlling the performance of rumen buffer materials is of significant interest to feed mills and end-user producers. The aim of this study was to evaluate, develop and optimise a laboratory protocol to consistently and robustly evaluate rumen buffering materials in order to predict their in vivo efficacy. Three different methods were evaluated for determining the buffering potential of carbonate buffer materials: (a) 2 and 8 h static pH, (b) 8 h fixed HCl acid load addition and (c) 3 h acidotic diet simulation using acetic acid. Buffer material, threshold pH, test duration and interactions between all three variables were significant (p < 0.001) in evaluating the performance of the buffer materials. The acidotic diet simulation was found to provide a different ranking of materials to the 8 h fixed HCl acid load methodology. The results highlight the importance of method selection and test parameters for accurately evaluating the potential efficacy of rumen buffer materials.