PubMed · 12695098
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James Beck. 2003. Visualizing neural networks.. https://doi.org/10.1016/s1046-2023(03)00002-1
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In recent years, both academia and pharmaceutical industry have produced significant advances in confocal detection and spectroscopy by laser-induced fluorescence. Confocal fluorescence studies provide information on identity, size, diffusion coefficient and concentration of the fluorescently labeled entity. This enables the establishment of sophisticated biochemical drug screening assays using the multitude of fluorescence parameters that can be observed (e.g. molecular brightness, fluorescence lifetime, anisotropy, resonance energy transfer). In cellular screening assays, confocality introduces spatial resolution in the vertical direction and reduces background fluorescence from outside the focal plane. Confocal HTS systems focusing on femtoliter-sized observation volumes allow for assay volumes far beyond current limits.
Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is a powerful analytical tool for the structural characterization of proteins and nucleic acids. However, many proteomics or genomics methodologies that employ MALDI-MS require external sample manipulation, which limits the overall throughput of analysis. We have focused on fabricating functional MALDI sample plates that would permit the on-probe characterization of nucleic acids. Here, we present results arising from the fabrication of functional sample plates composed of poly(methyl methacrylate) (PMMA). The PMMA sample plates were fabricated by a CNC milling technique. The key structural feature of our microfabricated samples plates is the presence of individual cylindrical posts (360 microm x 360 microm), which serve as individual sample targets within the overall PMMA-based MALDI sample plate. Functionality is added to these microposts via the covalent attachment of enzymes. As an example of the applicability of these microfabricated sample plates, enzymatic digestion of ribonucleic acids was performed on probe (i.e., on the micropost) with subsequent analysis by MALDI-MS. Advantages to such an approach include a reduction in sample handling (and concomitant sample losses) and a reduction in the amount of sample required for analysis due to the small surface area of the microposts.
We demonstrate, what is to the best our knowledge, a new method for studying the motion of a particle trapped by optical tweezers; in this method the trapping beam itself is used as a confocal probe. By studying the response of the particle to periodic motion of the tweezers, we obtain information about the medium viscosity, particle properties, and trap stiffness. We develop the mathematical model, demonstrate experimentally its validity for our system, and discuss advantages of using this method as a new form of scanning photonic force microscopy for applications in which a high spatial and temporal resolution of the medium viscosity is desired.