Label-free tetra-modal molecular imaging of living cells with CARS, SHG, THG and TSFG (coherent anti-Stokes Raman scattering, second harmonic generation, third harmonic generation and third-order sum frequency generation).

We have developed a new multimodal molecular imaging system that combines CARS (coherent anti-Stokes Raman scattering), SHG (second harmonic generation), THG (third harmonic generation) and multiplex TSFG (third-order sum frequency generation) using a subnanosecond white-light laser source. Molecular composition and their distribution in living cells are clearly visualized with different contrast enhancements through different mechanisms of CARS, SHG, THG and TSFG. A correlation image of CARS and TSF reveals that the TSF signal is generated predominantly from lipid droplets inside a cell as well as the peripheral cell wall.

[1]  Virginijus Barzda,et al.  Visualization of mitochondria in cardiomyocytes by simultaneous harmonic generation and fluorescence microscopy. , 2005, Optics express.

[2]  Dirk Roos,et al.  Intracellular Chemical Imaging of Heme-Containing Enzymes Involved in Innate Immunity Using Resonance Raman Microscopy , 2004 .

[3]  Hiro-o Hamaguchi,et al.  Supercontinuum dynamically visualizes a dividing single cell. , 2007, Analytical chemistry.

[4]  Ji-Xin Cheng,et al.  A multimodal platform for nonlinear optical microscopy and microspectroscopy. , 2009, Optics express.

[5]  William A Mohler,et al.  Three-dimensional high-resolution second-harmonic generation imaging of endogenous structural proteins in biological tissues. , 2002, Biophysical journal.

[6]  Vincent Couderc,et al.  Quantitative CARS molecular fingerprinting of single living cells with the use of the maximum entropy method. , 2010, Angewandte Chemie.

[7]  Hiro-o Hamaguchi,et al.  Molecular-level investigation of the structure, transformation, and bioactivity of single living fission yeast cells by time- and space-resolved Raman spectroscopy. , 2005, Biochemistry.

[8]  D Yelin,et al.  Laser scanning third-harmonic-generation microscopy in biology. , 1999, Optics express.

[9]  H. Winful,et al.  Physical origin of the Gouy phase shift. , 2001, Optics letters.

[10]  B. Tromberg,et al.  Imaging cells and extracellular matrix in vivo by using second-harmonic generation and two-photon excited fluorescence , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[11]  A. Fabre,et al.  Imaging lipid bodies in cells and tissues using third-harmonic generation microscopy , 2005, Nature Methods.

[12]  Ji-Xin Cheng,et al.  Multimodal nonlinear optical microscopy , 2011, Laser & photonics reviews.

[13]  W. Webb,et al.  Live tissue intrinsic emission microscopy using multiphoton-excited native fluorescence and second harmonic generation , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[14]  Hung-Che Chen,et al.  Integrated multiple multi-photon imaging and Raman spectroscopy for characterizing structure-constituent correlation of tissues. , 2008, Optics express.

[15]  S. Jaspersen,et al.  The budding yeast spindle pole body: structure, duplication, and function. , 2004, Annual review of cell and developmental biology.

[16]  Haw Yang,et al.  Three-dimensional chiral imaging by sum-frequency generation. , 2006, Journal of the American Chemical Society.

[17]  B. Byers,et al.  Behavior of spindles and spindle plaques in the cell cycle and conjugation of Saccharomyces cerevisiae , 1975, Journal of bacteriology.

[18]  Nektarios Tavernarakis,et al.  In vivo imaging of cellular structures in Caenorhabditis elegans by combined TPEF, SHG and THG microscopy , 2008, Journal of microscopy.

[19]  Vincent Couderc,et al.  Ultrabroadband multiplex CARS microspectroscopy and imaging using a subnanosecond supercontinuum light source in the deep near infrared. , 2008, Optics letters.

[20]  F. Ganikhanov,et al.  Multimodal nonlinear optical imaging of collagen arrays. , 2008, Journal of structural biology.

[21]  Massoud Motamedi,et al.  In vivo multimodal nonlinear optical imaging of mucosal tissue. , 2004, Optics express.

[22]  Ji-Xin Cheng,et al.  Green’s function formulation for third-harmonic generation microscopy , 2002 .