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There are two approaches for the investigation of light-tissue interactions: one numerical and the other analytical. The most common numerical method is the Monte Carlo (MC) simulation, which is a statistical study of photon migration from the optical properties of the different tissue regions. However, the yield of such statistical approaches is limited. Analytical methods are commonly based on the diffusion theory, yet they are inaccurate near the light source. There are several solution methods, where extrapolated boundary conditions lead to a more accurate solution.
Previously, we proposed measuring the full scattering profile (FSP), which is the angular intensity distribution, of cylindrical tissues. MC simulations revealed that the FSP has a fixed intensity point, named the iso-pathlength (IPL) point, which does not depend on the reduced scattering coefficient. The location of this point depends solely on the geometry. In this study, we derived the FSP from the steady state diffusion theory based on the extrapolated zeroboundary condition. A comparison of different reduced scattering coefficients revealed the IPL point, where the intensity remains constant. Furthermore, a MC simulation was performed under the same geometry and optical properties. We show that the position of the IPL according to the diffusion theory is in agreement with the MC simulation.
GNPs have a well-known effect on nearby fluorophores in terms of their fluorescence intensity (FI – increase or decrease) as well as fluorescence lifetime (FLT). We have designed a few bio-switch systems in which the FLIMdetected fluorescence varies after biologically relevant stimulation. Some of our tools include fluorescein diacetate (FDA) which can be activated by either esterases or pH, peptide chains cleavable by caspase 3, and the polymer polyacrylic acid which varies in size based on surrounding pH. After conjugating GNPs to chosen fluorophores, we have successfully demonstrated the logic gates of NOT, AND, OR, NAND, NOR, and XOR by imaging different stages of activation. These logic gates have been demonstrated both in solutions as well as within cultured cells, thereby possibly opening the door for nanoparticulate in vivo smart detection.
While these initial probes are mainly tools for intelligent detection systems, they lay the foundation for logic gates functioning in conjunction so as to lead to a form of in vivo biological computing, where the system would be able to release proper treatment options in specific situations without external influence.
Furthermore, we studied a group of patients without detection of inflammation that were sick. We found that this group was divided into two groups; one group had the same median FLT as the controls, and the other group had the same median FLT as the inflammatory patients. As a result, we believe the FD-FLIM system can suggest a faster and more accurate diagnostic technique than the methods used today. The correlations of the FLT distribution pattern with the different groups are presented.
Tracing apoptosis and stimulation in individual cells by fluorescence intensity and anisotropy decay
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During the past 20 years there has been a remarkable growth in the use of fluorescence in the biological sciences. Fluorescence spectroscopy and time-resolved fluorescence are considered to be primarily research tools in biochemistry and biophysics. This emphasis has changed, and the use of fluorescence has expanded. Fluorescence is now a dominant methodology used extensively in biotechnology, flow cytometry, medical diagnostics, DNA sequencing, forensics, and genetic analysis, to name a few.
The lectures will deal with basics of steady-state and time-resolved fluorescence spectroscopy, instrumentation and data analysis. They will cover time-domain and frequency-domain measurements, anisotropy, quenching and Förster Resonance Energy Transfer (FRET). Next, the lectures cover advanced time-resolved fluorescence topics and data analysis. Applications of fluorescence in biophysics, sensing, plasmon controlled fluorescence or material science are presented along with an introduction to fluorescence microscopy.
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