Raman spectroscopy: the gateway into tomorrow's virology

Abstract

In the molecular world, researchers act as detectives working hard to unravel the mysteries surrounding cells. One of the researchers' greatest tools in this endeavor has been Raman spectroscopy. Raman spectroscopy is a spectroscopic technique that measures the unique Raman spectra for every type of biological molecule. As such, Raman spectroscopy has the potential to provide scientists with a library of spectra that can be used to unravel the makeup of an unknown molecule. However, this technique is limited in that it is not able to manipulate particular structures without disturbing their unique environment. Recently, a novel technology that combines Raman spectroscopy with optical tweezers, termed Raman tweezers, evades this problem due to its ability to manipulate a sample without physical contact. As such, Raman tweezers has the potential to become an incredibly effective diagnostic tool for differentially distinguishing tissue, and therefore holds great promise in the field of virology for distinguishing between various virally infected cells. This review provides an introduction for a virologist into the world of spectroscopy and explores many of the potential applications of Raman tweezers in virology.

Figure 1

Line drawing depicting the region where Near UV and Near Infrared Wavelengths fall in the Light Spectrum.

Figure 2

Raman tweezers. The figure has been adapted from Hamden et al., 2005 [67]. The figure is a schematic of a model Raman tweezers. The combined laser tweezers and Raman spectroscopy instrument possesses a laser beam at 785 nm from a wavelength-stabilized, beam shape-circulated semiconductor diode laser that is introduced into an inverted microscope through a high numerical aperture objective (100×, NA = 1.30) to form an optical trap. The same laser beam is used to excite Raman scattering of the trapped particle. The scattering light from the particle is collected by the objective and coupled into a spectrograph through a 200-μm pinhole, which enables confocal detection and rejection of off-focusing Rayleigh scattering light. A holographic notch filter is used as a dichroic beam splitter that reflects the 785-nm excitation beam and transmits the Raman shifted light. A green-filtered illumination lamp and a video camera system are used to verify trapping and observe the image of the cell. The spectrograph is equipped with a liquid-nitrogen-cooled charge-coupled detector (CCD). Abbreviations: M-mirror; L-lens; DM-dichroic mirror; PH-pinhole; HNF-holograph notch filter.