Temporal Imaging of Live Cells by High-Speed Confocal Raman Microscopy.

Jeon Woong Kang,Niyom Lue,Freddy T Nguyen

doi:10.3390/ma14133732

Jeon Woong Kang, Niyom Lue + Show 1 more

Open Access

https://doi.org/10.3390/ma14133732

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Abstract

Label-free live cell imaging was performed using a custom-built high-speed confocal Raman microscopy system. For various cell types, cell-intrinsic Raman bands were monitored. The high-resolution temporal Raman images clearly delineated the intracellular distribution of biologically important molecules such as protein, lipid, and DNA. Furthermore, optical phase delay measured using quantitative phase microscopy shows similarity with the image reconstructed from the protein Raman peak. This reported work demonstrates that Raman imaging is a powerful label-free technique for studying various biomedical problems in vitro with minimal sample preparation and external perturbation to the cellular system.

Highlights

Since it was first observed in 1928 [1], Raman spectroscopy has been broadly used as an analytical technique in various fields
A Ti: Sapphire laser (3900S, Spectra-Physics, Milpitas, CA, USA) with 785 nm output wavelength was used as an excitation source of the custom-built NIR confocal Raman microscopy system (Figure 1) [11]
In that case, collecting as much Raman photons with a larger-core fiber is more relevant than collecting less signal while maintaining high resolution

Summary

Introduction

Since it was first observed in 1928 [1], Raman spectroscopy has been broadly used as an analytical technique in various fields. Scattered Raman photons from the sample includes the “finger print” information about the sample. By analyzing Raman photons, chemical composition of the sample can be obtained qualitatively and quantitatively. Confocal microscopy has been utilized to obtain 3-D structural information from cells [2]. The confocal pinhole only accepts the light from the focus resulting in high axial resolution. Confocal microscopy was implemented with reflectance, fluorescence, or even with inelastically scattered photons including Raman [3] or Brillouin [4]. Combining confocal microscopy and Raman spectroscopy provides the three-dimensional chemical mapping of biological samples with high spatial resolution

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