A wide‐field arrangement for single‐shot CARS imaging of living cells

Abstract

This paper demonstrates a new arrangement for wide-field CARS imaging. The setup using nanosecond dye lasers in a nonphase-matching illumination is presented. The chemical selectivity of the signal is demonstrated on polystyrene beads. Moreover, the experimental arrangement has enabled the acquisition of images of living cells in suspension with a satisfactory signal-to-background ratio by using only a single pair of laser shots. Therefore, each image can be acquired using a total duration of 3 ns, which is the pulse duration of the laser used in this study. Finally, lack of damage on cells has been demonstrated in conditions leading to strong CARS signal generation. In particular, no cell morphological changes were observed and cell membrane integrity (assessed by trypan blue test) was preserved.

Recent advances in linear and nonlinear Raman spectroscopy. Part VII

“…The optical setup uses nanosecond dye lasers in a nonphase‐matching illumination geometry. The experimental arrangement has enabled the acquisition of images of living cells in suspension with a satisfactory signal‐to‐background ratio by using only a single pair of laser shots . Walter and co‐workers described the Raman spectroscopic detection of nickel impact on single Streptomyces cells thereby revealing possible bioindicators for heavy metal contamination.…”

Advances in nonlinear optical spectroscopies: a historical perspective of developments and applications presented at ECONOS

“…A CARS microscopy method employing a wide‐field geometry was proposed by Heinrich et al Using a nanosecond laser source, the concept was demonstrated by fast imaging of oil droplets and sunflower seeds with chemical contrast. Another wide‐field CARS configuration was developed by Silve et al , who employed it for live imaging of cells.…”

Wide-field Surface-Enhanced Coherent Anti-Stokes Raman Scattering Microscopy

“…Early wide-field CARS setup used a dark-field condenser to focus pump light and counterpropagating Stokes beam to satisfy phase-matching condition. [24] Later, simper illumination geometries were adapted such as defocused laser beams [25] and high-incidence illumination schemes [26][27][28] . Such non-phase matched illuminations rely on the strongly scattering samples to redirect the light path.…”