Dark-Field Hyperspectral Imaging (DF-HSI) Modalities for Characterization of Single Molecule and Cellular Processes

Hyperspectral dark-field spectroscopy is achieved by combing a dark-field optical microscope with a hyperspectral detector. Due to recent advancements in optics, detection systems, image reconstruction, and computational capacity, the optical resolution of hyperspectral imaging (HSI) can now be translated down to micrometer and nanometer dimensions. Thus, making it possible to exquisitely detect and analyze single nanostructures to heterogeneous biological systems. HSI can now be used to validate, assist, and replace traditional imaging and spectroscopic technologies used to study plasmonic nanoparticles. Plasmonic nanostructures are of tremendous importance to study DNA coupling mechanisms in biosensors, virus–host interactions in the infected cells, live cell imaging coupled with motion detection of nano-objects, cancer detection, immunotherapies, toxicity and transport mechanisms, and targeted drug delivery. Many biological problems can be solved by capturing and understanding electromagnetic radiation–matter interactions in the form of scattering, transmission, and absorption by a high-throughput and robust imaging modality. Another active area of research where HSI has been used as a low-cost, label-free technique is to investigate single cellular processes, single molecule interactions, and un-mix information of tissue-level structures in the surgical procedures. This book chapter provides a brief description attributed to working principle of dark-field hyperspectral imaging. Further, advanced applications of HSI in detecting single and ensemble plasmonic nanostructures, single molecule events, environmental sensing, single cell imaging, biological analysis, and surgical vision are described.