Label-free cellular imaging by broadband coherent anti-Stokes Raman scattering microscopy.
Raman microspectroscopy can provide the chemical contrast needed to characterize the complex intracellular environment and macromolecular organization in cells without exogenous labels. It has shown a remarkable ability to detect chemical changes underlying cell differentiation and pathology-related chemical changes in tissues but has not been widely adopted for imaging, largely due to low signal levels. Broadband coherent anti-Stokes Raman scattering (B-CARS) offers the same inherent chemical contrast as spontaneous Raman but with increased acquisition rates. To date, however, only spectrally resolved signals from the strong CH-related vibrations have been used for CARS imaging. Here, we obtain Raman spectral images of single cells with a spectral range of 600-3200 cm⁻¹, including signatures from weakly scattering modes as well as CH vibrations. We also show that B-CARS imaging can be used to measure spectral signatures of individual cells at least fivefold faster than spontaneous Raman microspectroscopy and can be used to generate maps of biochemical species in cells. This improved spectral range and signal intensity opens the door for more widespread use of vibrational spectroscopic imaging in biology and clinical diagnostics.
Polymers Division, National Institute of Standards and Technology, Gaithersburg, Maryland, USA.
Sapun H
SH
Parekh
Young Jong
YJ
Lee
Khaled A
KA
Aamer
Marcus T
MT
Cicerone
D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D051379:Mice; D008853:Microscopy / Q000379:methods*; D057054:Molecular Imaging / Q000379:methods*; D059010:Single-Cell Analysis; D013059:Spectrum Analysis, Raman / Q000379:methods*; D014732:Vibration