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Novel optical detection methods in b...

University of California, Berkeley.

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Novel optical detection methods in biophysics.

Record Type:	Language materials, printed : Monograph/item
Title/Author:	Novel optical detection methods in biophysics./
Author:	Siu, Merek Columba.
Description:	117 p.
Notes:	Advisers: Jan T. Liphardt; Carlos J. Bustamante.
Contained By:	Dissertation Abstracts International69-03B.
Subject:	Biophysics, General. -
Online resource:	http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=3306347
ISBN:	9780549529538

Novel optical detection methods in biophysics.
Siu, Merek Columba.

Novel optical detection methods in biophysics. - 117 p.

Advisers: Jan T. Liphardt; Carlos J. Bustamante.

Thesis (Ph.D.)--University of California, Berkeley, 2007.

The cell can be described as a coordinated factory where many functions are accomplished by molecular machines. To understand how individual machines work, and work together, tools that measure relative motions, the forces that cause these movements, and the precise location of a given object over time are invaluable. Conventional optical microscopies can resolve distance changes on scales beyond the diffraction limit of &sim;250 nm, or on the 1--10 nm scale accessible by Fluorescence Resonance Energy Transfer (FRET). The intermediate length regime of 10--100 nm can now be addressed by plasmon rulers. The readout for this technique is the distance dependent change in the wavelength and intensity of light scattered off gold or silver nanoparticle dimers. Since this method is based on light scattering instead of fluorescence from the organic dyes typically used in FRET, essentially unlimited observation times under continuous illumination are possible. Here, we establish an empirical calibration of the plasmon rulers that establishes the plasmon resonance wavelength versus distance relationship under experimentally realistic situations.

ISBN: 9780549529538Subjects--Topical Terms:

1019105
Biophysics, General.

Novel optical detection methods in biophysics.
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020 $a 9780549529538
035 $a (UMI)AAI3306347
035 $a AAI3306347
040 $a UMI $c UMI
100 1 $a Siu, Merek Columba. $3 1019108
245 1 0 $a Novel optical detection methods in biophysics.
300 $a 117 p.
500 $a Advisers: Jan T. Liphardt; Carlos J. Bustamante.
500 $a Source: Dissertation Abstracts International, Volume: 69-03, Section: B, page: 1513.
502 $a Thesis (Ph.D.)--University of California, Berkeley, 2007.
520 $a The cell can be described as a coordinated factory where many functions are accomplished by molecular machines. To understand how individual machines work, and work together, tools that measure relative motions, the forces that cause these movements, and the precise location of a given object over time are invaluable. Conventional optical microscopies can resolve distance changes on scales beyond the diffraction limit of &sim;250 nm, or on the 1--10 nm scale accessible by Fluorescence Resonance Energy Transfer (FRET). The intermediate length regime of 10--100 nm can now be addressed by plasmon rulers. The readout for this technique is the distance dependent change in the wavelength and intensity of light scattered off gold or silver nanoparticle dimers. Since this method is based on light scattering instead of fluorescence from the organic dyes typically used in FRET, essentially unlimited observation times under continuous illumination are possible. Here, we establish an empirical calibration of the plasmon rulers that establishes the plasmon resonance wavelength versus distance relationship under experimentally realistic situations.
520 $a Forces, which are responsible for distance changes, are also key to understanding molecular machines. Mechanical manipulation technologies such as optical tweezers, atomic force microscopy, and calibrated microneedles, can precisely measure and exert forces on the picoNewton to nanoNewton range. However, the general requirement for direct physical attachment to micron-sized handles makes in vivo measurements challenging. Additionally, these techniques are best suited for measuring externally applied tensile forces instead of forces that develop internal to macromolecules. Optical force sensors, comprised of a single-stranded DNA spring whose extension is read out by FRET, can address some of these limitations. We describe an application of optical force sensors to exploring the interplay between DNA internal force, DNA sequence, and protein-DNA binding affinity. We studied the binding of Integration Host Factor (IHF), a bacterial protein that sharply bends DNA, to constructs strained by a variety of internal forces. Less than 3 pN of internal force is sufficient to increase IHF binding affinity tenfold. Furthermore, internal force can completely rescue binding affinity to mutant DNA constructs that exhibit significantly reduced binding affinities in their unstrained forms.
520 $a Complementing distance change and force measurements, the evolution of spatial position over time can yield mechanistic information about molecular machines. Here, we describe work towards tracking single large cargos as they transport through the nuclear pore complex. A model large cargo consisting of a quantum dot coupled to an import signal from Snurportin I has been validated. High resolution localization should reveal detailed cargo trajectories. The resulting movie of the translocation intermediate could help clarify how nuclear pores distinguish bona fide cargos from inert materials.
590 $a School code: 0028.
650 4 $a Biophysics, General. $3 1019105
690 $a 0786
710 2 $a University of California, Berkeley. $3 687832
773 0 $t Dissertation Abstracts International $g 69-03B.
790 $a 0028
790 1 0 $a Bustamante, Carlos J., $e advisor
790 1 0 $a Liphardt, Jan T., $e advisor
791 $a Ph.D.
792 $a 2007
856 4 0 $u http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=3306347