Strengths and limitations of size exclusion chromatography for investigating single chain folding – current status and future perspectives
Synthetic approaches for Single-Chain Nanoparticles (SCNPs) developed rapidly during the last decade, opening a multitude of avenues for the design of functional macromolecular chains able to collapse into defined nanoparticles. SCNPs aim at mimicking the behavior of natural biomacromolecules and are promising structures for catalysis or biomedicine, and thus need to be prepared with high synthetic fidelity, requiring precise analytical tools. One of the most frequently used methods for studying the formation of single-chain nanoparticles is Size Exclusion Chromatography (SEC). Despite its outstanding accessibility and flexibility, SEC is a relative method that – if not hyphenated to adequate detectors – leaves room for considerable misinterpretation. Herein, the limits of SEC for SCNP analysis are critically discussed. Furthermore, the need for appropriate auxiliary and complementary analysis techniques is highlighted.