Applications of coherent Raman scattering microscopy in bioimaging
Much progress in biology and allied sciences is driven by optical microscopy methods. Techniques involving extraneous labels (e.g. fluorescent molecules) with affinity to one or more sample components are widely used, with some implementations even breaching the diffraction limit. Other than the fact that the introduction of these labels changes sample chemistry, availability of suitable labels, issues of photobleaching, and phototoxicity can limit the applicability of these methods.
Spectroscopy techniques, which rely on energy-level transitions of various molecular species in the sample are chemically specific, sensitive, and label-free. Coherent Raman scattering (CRS), namely coherent anti-Stokes Raman scattering (CARS) and stimulated Raman scattering (SRS) have been applied in bioimaging, with short acquisition times and promising results.
In this talk, I will discuss my research at Cardiff University (PhD) and KU Leuven, where I used CARS and SRS microscopy to image biological samples. Using volumetric hyperspectral CARS microscopy [1-3], I studied mitosis in human osteosarcoma (U-2OS) cells, revealing the sub-cellular distribution of various biomolecules. As an outlook to pharmacodynamical studies, I imaged the effects of two anti-cancer drugs in single cells. In addition to these, I will also show some results from an exploratory application of CARS to 3D tissue cultures- organoids. Not only a proof of principle, this work has revealed interesting information previously unresolved with CRS.
As a classic example of how fluorescent molecules are not always the best choice for bioimaging, I will present results of SRS imaging of lipids in desert locust (Schistocerca gregaria) oocytes which do not stain efficiently using conventional methods.
When applied to statistically significant sample sizes, the methods demonstrated in this research could establish CARS and SRS as novel and extremely useful tools in biomedical/biochemical research.
1.Pope I, Langbein W, Watson P, Borri P, Optics Express 21, 7096 (2013)
2.Masia F, Glen A, Stephens P, Borri P, Langbein W, Anal. Chem. 85, 10820 (2013)
3.Karuna A, Masia F, Borri P, Langbein W, Journ. Raman Spec. 47, 1167 (2016)