During cell division, various organelles behave dynamically. Visualization of these dynamic behaviors of organelles is a promising one step forward for understanding life at the molecular level. One- or two-photon excited fluorescence microscopy has so far been used for visualizing these cell dynamics. The fluorescent probe introduced into a living cell can visualize the spatial distribution of a target molecule in real time, enabling the tracing of cell dynamics at the molecular level. Introducing a fluorescent probe into a cell, however, may alter the physical and chemical conditions of the cell. Here we show a new method for direct (no need for staining cells) visualization of living cell processes with coherent anti-Stokes Raman scattering (CARS) spectroscopy. A new light source, supercontinuum generated from a photonic crystal fiber, has facilitated ultrabroadband (>3500 cm-1) multiplex CARS spectroscopy and imaging with high molecular specificity. Using this multiplex CARS technique, we have been successful in tracing the whole cell division process, the splitting of a mother cell into two daughter cells, appearance and disappearance of septum, and dynamic distribution changes of organelles consisting of lipid membrane. The supercontinuum has also facilitated simultaneous measurement of the CARS and two-photon excited fluorescence (TPEF) spectra, enabling what we call multiple nonlinear spectral imaging. Three-dimensional image reconstruction of a living cell with high speed is now possible to elucidate more detailed molecular-level dynamics inside a dividing living cell.
All Science Journal Classification (ASJC) codes
- Analytical Chemistry