Specimen preparation optimization for size and morphology characterization of nanocellulose by TEM

Nanoparticle morphology, size and dispersion are key parameters for the application of cellulose nanomaterials in various areas, such as polymer nanocomposites, catalysts, gel and so on. Transmission electron microscopy (TEM) is the most suitable technique for the morphological characterization of these particles. However, nanocellulose low contrast in TEM images is the major drawback for their adequate morphological characterization and size determination. Even though it is widespread knowledge that negative staining using uranyl acetate is the best approach for intensifying cellulose contrast, up to now few have succeeded in achieving high quality images and reliable size measurements of these nanomaterials. This protocol presents an optimization of the standard uranyl acetate protocol commonly used for biological specimens in order to suit cellulose nanomaterials. Drying method and grid conditions were proven to be the most significant variables for effective TEM specimen preparation. These guidelines could also be successfully applied to enhance the cellulose nanomaterial contrast in polymer matrices.