Bench-scale methods such as Biochemical Methane Potential (BMP) assays and Anaerobic Toxicity Assays (ATAs) are useful tools in evaluating potential feedstocks for anaerobic digestion. The BMP method provides a preliminary indication of substrate biodegradability and methane production, while the ATAs provide an indication of substrate toxicity to anaerobic microbial consortia. Previous research using small (<20 >L) reactors indicated that co-digestion of manures with small amounts of glycerin (ca. 1 – 2 %) can double methane production, but toxicity can result if glycerin exceeds 2% (volumetric basis). This paper investigated the relationship between bench-scale methods (BMPs and ATAs) and sub pilot-scale digester results, using glycerin as a test substrate mixed with a baseline feedstock (beef manure, corn processing wastewater, lagoon liquid, and short-fiber cardboard). The batch-fed, stirred ATAs indicated that glycerin was toxic to methane production at all inclusion levels. The batch-fed, stirred BMPs indicated no significant difference between methane production in the 0.0%to 4.0% addition levels; however at 8.0% addition, methane production tripled. The continuously fed, non-stirred, plug-flow sub pilot-scale reactors indicated toxicity effects in the 2.0% and 4.0% glycerin mixtures and no difference from the control in the 1.0% glycerin mixture. These results demonstrate the variations in scale performance of glycerin as a co-substrate and identify some serious challenges in extrapolating bench-scale assays to large-scale performance of mixed-waste anaerobic digestion systems.