Traditional glass or polymer-based substrates do not participate in the enhancement of the cellular imaging process. In fact, more than 80% of the total incident light is lost into the substrates. Existing methods for improving imaging sensitivity such as Total Internal Reflection Fluorescence Microscopy and metal-coated slides provide planar fluorescence enhancement by using evanescent waves to excite a spatially confined zone. The Illinois-based team on the other hand, decided to use a nanostructured surface, made on a silicon based substrate using a conventional plasma-based etching method, to enhance cell imaging.

When Chinese Hamster Ovary cells, stained with fluorescence dyes, were imaged on the nanostructured surface, three-dimensional fluorescence enhancement was observed. The detailed optical analysis of the substrate showed the possibility of coupling incident light from the nanostructured surface into the cells. Due to enhanced scattering from the surface coupled with multiple internal reflections bounded by the cell membrane, light trapping can take place inside the cell, thus allowing higher fluorescence intensity. The same enhancement can also be observed if the cells are transfected with fluorescent protein.

Fireworks inside cells

The group's results provide a new way to achieve higher sensitivity during cell imaging. As opposed to two-dimensional near field enhancement in existing methods, three-dimensional far field enhancements can be obtained with the surface plasmon-based substrate. There is the possibility of tuning the surface plasmon resonance so that a specific fluorescence molecule or protein gets enhanced inside the cells. This may offer the 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.

The researchers presented their research in the journal Nanotechnology.