Label-Free Techniques for Probing Biomolecular Condensates

Biomolecular condensates play important roles in a wide array of fundamental biological processes, such as cellular compartmentalization, cellular regulation, and other biochemical reactions. Since their discovery and first observations, an extensive and expansive library of tools has been developed to investigate various aspects and properties, encompassing structural and compositional information, material properties, and their evolution throughout the life cycle from formation to eventual dissolution. This Review presents an overview of the expanded set of tools and methods that researchers use to probe the properties of biomolecular condensates across diverse scales of length, concentration, stiffness, and time. In particular, we review recent years' exciting development of label-free techniques and methodologies. We broadly organize the set of tools into 3 categories: (1) imaging-based techniques, such as transmitted-light microscopy (TLM) and Brillouin microscopy (BM), (2) force spectroscopy techniques, such as atomic force microscopy (AFM) and the optical tweezer (OT), and (3) microfluidic platforms and emerging technologies. We point out the tools' key opportunities, challenges, and future perspectives and analyze their correlative potential as well as compatibility with other techniques. Additionally, we review emerging techniques, namely, differential dynamic microscopy (DDM) and interferometric scattering microscopy (iSCAT), that have huge potential for future applications in studying biomolecular condensates. Finally, we highlight how some of these techniques can be translated for diagnostics and therapy purposes. We hope this Review serves as a useful guide for new researchers in this field and aids in advancing the development of new biophysical tools to study biomolecular condensates.