High Resolution Light Microscopy
Fluorescence microscopy has experienced tremendous growth over the past four decades of the century. It also has facilitated major advancements in the fields of science. The classic fluorescent techniques, including epi-fluorescence and confocal, has allowed researchers to selectively observe labeled structures with great clarity and consistency. Historically, biological scientists have been the most prolific users of fluorescence imaging. However, the increasing numbers of nano-materials research initiatives are now incorporating quantum dots and other fluorescent labels into their imaging protocols. This is especially true in areas where nano-materials and biological research is overlapping. Example, in cases like drug delivery. This nano-bio convergence, along with other advancements, has generated the need to observe highly dynamic events involving labeled and unlabeled structures in real time. For many researchers, observing the sample portion that is fluorescently labeled is often not sufficient.
Many times it is important to visualize the fluorophore labeled components in the full context of the sample. The traditional methods of widefield epi-fluorescence and confocal microscopy are highly effective techniques for quantifying that portion of the sample that is fluorescent. However, in order to visualize these fluorescent components in the full context of the sample, additional imaging techniques such as differential interference contrast, or DIG, and phase contrast are often needed and utilized. Traditionally, these techniques are used to capture the refractive structure of the sample. These images are combined with the fluorescent image. This is commonly referred to as an the overlay technique.
The Dual Mode Fluorescence, or DMF, module was recently added to the high resolution capability of CytoViva. DMF is a transmitted light fluorescence imaging technique that allows both fluorescently labeled and non-labeled sample structures to be observed simultaneously and in real time. Using some multiple filter configurations, a wide variety of fluorophores can be observed with the use of this technique. This can include specialized filter sets optimized for quantum dot observation. Both excitation and emission filters used in this technique are available from major filter manufacturers. The DMF technique uses a classic mercury halide illumination source with spectral peaks optimized for fluorescence. The light source, DMF module and high resolution adapter are connected with the liquid light guides. This modular configuration allows the technique to be utilized on a wide range of upright and inverted light microscopes. The convergence of nano-materials and biological science has become significant for both academic and industrial research communities. Applications that utilize nano-materials to manipulate and interact with biological specimens hold great promise. This also represents one of the fastest growing segments of basic research. This Nano-bio related research is also driving the need for fluorescence microscopy techniques. Nano-materials have already moved out of the lab and are being incorporated into our everyday lives. For example, silver nano-particles have been integrated into clothing and other materials due to their anti-microbial properties. Furthermore, for scientists involved in nano-materials development, light microscopy is often viewed as a complimentary tool to SEM, TEM and AFM systems. For example, research conducted by NASA recently noted that optical microscopy is an important tool for initial assessments of single wall nanotubes to evaluate the dispersions of aggregates before moving on to higher resolution microscopy methods.
This technique also has its important biological applications. Biological sciences have long been the primary domain of fluorescence-based microscopy due to the high level of specificity available through immunocytochemistry. However the increasing need to observe live cells, combined with an ever-expanding number of fluorescent reagents and transgenic fluorescent proteins increases the demand for a wide range of fluorescent imaging techniques. The use of CytoViva DMF allow the researcher to maximize the advantages of many of the physic reagents and modern biological fluorescent techniques by allowing for real time observation of fluorescence in the full context in the sample environment. Not only does this technique provide time context of the fluorophore labels within the sample, interactions between these sample components can also be easily observed.