Live Cell imaging of Host-Virus interactions using CARS and Two-Photon microscopy

Confocal microscopy is an established imaging technique, however the addition of exogenous labels can disrupt cellular functions and often limits the technique to studying fixed samples. In contrast, coherent anti-Stokes Raman scattering (CARS) microscopy enables non-invasive intracellular imaging without the need for labelling. The CARS technique has been applied to a variety of biological specimens due to its properties of enhanced signal and chemical selectivity, enabling intracellular imaging with vibrational contrast at high acquisition speeds. In this way, CARS provides the capability for live cell imaging.

We have developed a multi-modal microscope platform that combines CARS and two-photon fluorescence microscopies in order to study the effects of viral infection on host cell morphology and lipid droplet distrubution. We apply this to study living fibroblast cells infected with cytomegalovirus. The virus is genetically modified to cause expression of the green fluorescent protein (GFP) in the host cell upon infection. The CARS laser source is tuned to the C?H stretch vibration at 2845 cm-1 in order to visualise the cell morphology and lipid droplets. The two-photon images reveal the extent of viral protein expression due to the amount of GFP expressed.

Therefore, using this microscope platform, we can show host-pathogen interactions by monitoring cellular changes, such as changes in lipid droplet distribution and nucleus morphology during live infection. This method has potential applications in the study of other host-virus interactions as well as the capability to study changes of cellular material other than lipid droplets.