Enzyme penetration into plant cell wall is widely acknowledged to be a key bar­rier to economical and effective biochemical conversion of lignocellulosic biomass [5, 49]. In fact, the primary function of pretreatment of lignocellulosic biomass is to assist subsequent enzymatic digestibility by making cell walls more accessi­ble to saccharifying enzymes [1, 4, 44]. However, an accurate description of the methods by which enzymes penetrate cell walls and accomplish cellulose degra­dation has been lacking. A recent study by Donohoe and coworkers provided, for the first time, direct visual evidence of loosening of plant cell wall structure due to dilute acid pretreatment and the subsequently improved access by cellulases [49]. Figure 3c-f further demonstrate the penetration of cellulases into pretreated cell walls as detected by nano-gold labeled antibodies to Cel7A and other cellulases. This study shows that penetration of enzymes into mildly pretreated cell walls is minimal and that cells stay largely intact even after prolonged exposure to cellu­lases (Fig. 3a, b). In moderately pretreated cell walls, cellulases are able to partially penetrate and disintegrate the inner secondary layers (S3) only (Fig. 3c, d); whereas the outer layers (S1 and S2) remain impervious to enzymes. In severely pretreated cell walls, enzymes penetrate throughout (Fig. 3e, f). These data suggest that enzy­matic digestibility of biomass is restricted by transport of enzymes into cell walls. While not directly evidenced by this study, these results also suggest that thermal pretreatments (and possibly others) “loosen” cell walls in layers providing enzymes access only to these structurally compromised zones of the cell walls. Kinetic data on thermal pretreatments by several research groups also suggests likely mass trans­fer limited xylan removal that can be modeled as parallel fast and slow reactions [44, 50, 51] and the fundamental observations made by Donohoe and coworkers [49] support this hypothesis.