A CARS solution with high temporal resolution

Abstract

Confocal and multiphoton microscopy are powerful fluorescence techniques for morphological and dynamics studies of labeled elements. For non-fluorescent components, CARS (Coherent Anti-Stokes Raman Scattering) microscopy can be used for imaging various elements of cells such as lipids, proteins, DNA, etc. This technique is based on the intrinsic vibrational properties of the molecules. Leica Microsystems has combined CARS technology with its TCS SP5 II confocal microscope to provide several advantages for CARS imaging. The Leica TCS SP5 II combines two technologies in one system: a conventional scanner for maximum resolution and a resonant scanner for high time resolution. For CARS microscopy, two picosecond near-infrared lasers are tightly overlapped spatially and temporally and sent directly into the confocal system. The conventional scanner can be used for morphological studies and the resonant scanner for following dynamic processes of unstained living cells. The fast scanner has several advantages over other solutions. First, the sectioning is truly confocal and does not suffer from spatial leakage. Second, the high speed (29 images/sec @ 512x512 pixels) provides fast data acquisition at video rates, allowing studies at the sub-cellular level. In summary, CARS microscopy combined with the tandem scanner makes the Leica TCS SP5 II a powerful tool for multi-modal and three-dimensional imaging of chemical and biological samples. We will present our solution and show results from recent studies with the Leica instrument to illustrate the high flexibility of our system.