A major factor hindering the economic viability of biomass utilization is the high cost of enzymatic deconstruction of cellulose. To attain substantial improvements, detailed molecular-level understanding of the interaction between cellulosic substrates and cellulases is needed. The effect of the structure of cellulose, cellulose surface topography and crystallinity, as well as the chemistry of the local environment on cellulase binding and enzyme synergism need to be elucidated in order to engineer optimized pretreatment and reaction conditions. Within that aim, we have developed a versatile platform that incorporates atomic force microscopy (AFM), confocal microscopy and biochemical assays to fully characterize cellulose deconstruction by cellulases at relevant scales. Changes in cellulose structure with sub-nanometer resolution along with enzyme binding were observed in an ongoing hydrolysis reaction with up to 90% hydrolysis.  Additionally, simultaneous confocal microscopy was used to locate fluorophore labeled cellulose microfibrils or bound fluorophore- labeled enzyme. The system requires minimal sample preparation and allows monitoring of a hydrolysis reaction for extended time periods in the AFM stage. We believe this platform can be extended to investigate the deconstruction of cell wall models in which confocal microscopy will further serve to co-locate different cell wall components.