Aqueous ammonia pretreatment was optimized and the limiting factors in high solids enzymatic hydrolysis were assessed. The recommended pretreatment condition to achieve high enzymatic yield was: 180°C, 20% (w/w) ammonia, 30min, and 20% solids content. FT-IR and GC-MS results indicated that most of the lignin was degraded to soluble fragments after pretreatment. The pretreated solids after post-washing showed higher enzymatic digestibility at high solids loading than that without washing. The washed solids required lower cellulase and xylanase dosage than unwashed solids to achieve high sugar yield. Enzymatic conversions were declined with the increased solids loading of pretreated solids, pretreated-washed solids, and filter papers. The results indicated that solids loading in enzymatic hydrolysis was an important factor affecting sugar yield. The increasing concentration of glucose and ligno-phenolics mainly inhibited the enzymatic hydrolysis of aqueous ammonia pretreated corn stover.