Correlative Atomic Force Microscopy and Photo-Activated Localization Microscopy of Nano-cellulose Fibrils from Tunicate and Poplar*

The molecular structure of cellulose in its native nano-fibril form is difficult to characterize and consequently still not well understood. We report simultaneous Atomic Force Microscopy (AFM) and Photo-Activated Localization Microscopy (PALM) of cellulose nano-fibrils (CNFs) from Tunicate and Poplar. PeakForce Quantitative Nanomechanical Mapping (PFQNM) is employed to characterize sample stiffness and modulus mapping. To complement the AFM measurements, the nano-fibrils were exposed to recombinant family 3 Carbohydrate Binding Modules (CBM3) specifically binding to the hydrophobic surface of crystalline CNF. The CBM3 was also tagged with a photoactivatable fluorescent protein PA-mCherry to allow super resolution imaging by PALM. We compare the distribution of CBM binding sites to the nano-fibril topography to draw conclusions about the degree of crystallinity of the elemental CNF and the distribution of amorphous areas within the CNF and at morphological kinks, which show a non-Gaussian distribution related to the crystallographic planes of cellulose.