Dilute acid pretreated biomass is normally toxic and requires a conditioning step to remove compounds that inhibit fermentation of sugars to ethanol. The objective of this study was to evaluate the effectiveness of lime and ammonium hydroxide as conditioning agents for dilute acid pretreated corn stover. Pretreated stover was produced in a pilot-scale reactor and liquor extracted from the pretreated slurry was treated using either an “overliming” process or with ammonium hydroxide at various concentrations and treatment temperatures. The conditioned liquor was then spiked with additional glucose representing glucose that would be produced by enzymatic cellulose hydrolysis of the pretreated solids were the solids still present. Glucose and xylose in the treated liquor were fermented to ethanol using a recombinant Zymomonas mobilis 8b. A shake flask screening study identified conditions where treatment with ammonium hydroxide did not produce sugar losses normally seen during lime treatments. These results were confirmed in 0.5 L working volume fermentations in pH controlled fermentors, in which, 15% to 20% improvement in ethanol yields for ammonium hydroxide treated liquor compared to lime treated material was demonstrated and validated by a rigorous analysis of the mass balance closure data. In large measure, these improved results were achieved because the ammonium hydroxide treatment did not result in significant sugar losses.