Traditional glass or polymer based substrate does not participate in the enhancement of the cellular imaging process. In fact, more than 80% of the total incident light is lost in to the substrates. Existing methods to improve the imaging sensitivity such as Total Internal Reflection Fluorescence Microscopy and metal coated slides provide planar fluorescence enhancement by using evanescent wave to excite a spatially confined zone. We, on the other hand, decided to use nanostructured surface, made on silicon based substrate using conventional plasma based etching method, as the cell imaging surface. When Chinese Hamster Ovary cells, stained with fluorescence dyes, were imaged on the nanostructured surface, three-dimensional fluorescence enhancements was observed for the cells. The detailed optical analysis of the substrate showed the possibility of coupling incident light from the nanostructured surface in to the cells. Due to enhanced scattering from the surface coupled with multiple random reflections inside cells, light trapping can happen in the cell, thus allowing higher fluorescence intensity. The same enhancement can also be observed if the cells are transfected with fluorescent protein.
Our group's results provide a new way to achieve higher sensitivity during cell imaging. As oppose to one dimensional enhancement in existing methods, three dimensional enhancements can be obtained with the surface plasmon based substrate. There is possibility of tuning the surface plasmon resonance so that specific fluorescence molecule or protein gets enhanced inside the cells. This may offer opportunity to capture single molecular events in living cells, observe very early stage protein expressions soon after genetic transfection, and discover new basic mechanisms in cytoskeletal dynamics, cell adhesion and migration as well as membrane trafficking.
M. R. Gartia et al. Nanotechnology(2011).