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Analysis of Protein Aggregation in Sub-Femtoliter Volume by Two-Photon-Based Fluorescence Correlation Spectroscopy

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We presented a fluorescence correlation spectroscopy (FCS) based on two-photon microscopic system to identify physical parameters that give rise to fluctuations in fluorescence signal. The excitation volume is a function of the overall instrumental setting, that include the utilized laser power, the laser wavelength, the magnification, and the power of the objectives and other optics for the microscope. For a given excitation volume, the diffusion coefficient of molecules due to Brownian Movement into or out of this excitation volume can be determined. Furthermore, based on the autocorrelation analysis that is dependent on the diffusion coefficients of molecules, the fraction of bound species from measured samples containing the dye labeled anti-rabbit IgG and rabbit IgG can be separated. The mean standard deviation of the diffusion coefficients measured in our two-photon-based system is about 0.064. Moreover, the maximal binding fraction of kinetics mode and end mode between anti-rabbit IgG and rabbit IgG is 60% and 73%, respectively.
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Keywords: BROWNIAN MOVEMENT; DIFFUSION COEFFICIENT; EXCITAION VOLUME; FLUORESCENCE CORRELATION SPECTROSCOPY; TWO-PHOTON MICROSCOPY

Document Type: Research Article

Publication date: December 1, 2008

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  • Bionanoscience attempts to harness various functions of biological macromolecules and integrate them with engineering for technological applications. It is based on a bottom-up approach and encompasses structural biology, biomacromolecular engineering, material science, and engineering, extending the horizon of material science. The journal aims at publication of (i) Letters (ii) Reviews (3) Concepts (4) Rapid communications (5) Research papers (6) Book reviews (7) Conference announcements in the interface between chemistry, physics, biology, material science, and technology. The use of biological macromolecules as sensors, biomaterials, information storage devices, biomolecular arrays, molecular machines is significantly increasing. The traditional disciplines of chemistry, physics, and biology are overlapping and coalescing with nanoscale science and technology. Currently research in this area is scattered in different journals and this journal seeks to bring them under a single umbrella to ensure highest quality peer-reviewed research for rapid dissemination in areas that are in the forefront of science and technology which is witnessing phenomenal and accelerated growth.
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