Cotton gin waste was collected from a cotton ginning plant in Virginia and characterized before and after steam explosion to evaluate its potential applications for higher value products such as ethanol. The raw cotton gin waste had high levels of ash (10.5 wt%) and acid insoluble material (28.8 wt%). The xylan and cellulose contents were, respectively, 9 and 37 wt%. The cotton gin waste was steam exploded in a batch gun at severities ranging from 2 to 4.9. Substantial solubilization/degradation of fiber material (9-17 wt%) occurred during steam explosion pretreatment especially at the high severities. Mannan, galactan, and arabinan were completely solubilized/degraded at severities greater than 3.56. Both glucan and xylan were solubilized/degraded at all severities, but xylan loss was considerably higher. The steam exploded material was essentially cellulose and acid insoluble material and small fractions of xylan at severities greater than 3.56. Steam explosion improved the enzyme hydrolysis of the material from 42% to 67% during 24 h incubation. E. coli KO11 was effective in converting the enzyme hydrolyzed substrate to ethanol. The highest ethanol yield (83% of theoretical yield) was achieved for material treated at a severity of 3.56. Both xylose and glucose were fermented to ethanol by the microorganism.