Methodological guidelines for the characterization of polysaccharide helical structures

Polysaccharide chain conformation is a critical determinant of their physicochemical and biological properties; however, reliable identification remains a significant challenge due to the inconsistent structural evidence yielded by conventional analytical techniques. This guide presents a practical framework for characterizing the helical structures of polysaccharides. It systematically evaluates and compares the operating procedures of the Congo red spectral shift assay, atomic force microscopy (AFM), and light scattering (SLS/DLS), supplemented by auxiliary methods such as circular dichroism (CD) and transmission electron microscopy (TEM). Furthermore, beyond detailing fundamental principles, this work emphasizes critical experimental precautions to avoid analytical misinterpretations. This integrated approach provides a rational basis for method selection, enabling researchers to combine qualitative and quantitative techniques to obtain reliable evidence of triple‐helical conformations for subsequent structure‐function analyses.