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Interference techniques in fluoresce...
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Boston University.
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Interference techniques in fluorescence microscopy.
紀錄類型:
書目-電子資源 : Monograph/item
正題名/作者:
Interference techniques in fluorescence microscopy./
作者:
Dogan, Mehmet.
面頁冊數:
132 p.
附註:
Adviser: Bennett B. Goldberg.
Contained By:
Dissertation Abstracts International70-01B.
標題:
Biology, Cell. -
電子資源:
http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=3345716
ISBN:
9781109003154
Interference techniques in fluorescence microscopy.
Dogan, Mehmet.
Interference techniques in fluorescence microscopy.
- 132 p.
Adviser: Bennett B. Goldberg.
Thesis (Ph.D.)--Boston University, 2009.
We developed a set of interference-based optical microscopy techniques to study biological structures through nanometer-scale axial localization of fluorescent biomarkers.
ISBN: 9781109003154Subjects--Topical Terms:
1017686
Biology, Cell.
Interference techniques in fluorescence microscopy.
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Spectral self-interference fluorescence microscopy (SSFM) utilizes interference of direct and reflected waves emitted from fluorescent molecules in the vicinity of planar reflectors to reveal the axial position of the molecules. A comprehensive calculation algorithm based on Green's function formalism is presented to verify the validity of approximations used in a far-field approach that describes the emission of fluorescent markers near interfaces. Using the validated model, theoretical limits of axial localization were determined with emphasis given to numerical aperture (NA) dependence of localization uncertainty. SSFM was experimentally demonstrated in conformational analysis of nucleoproteins. In particular, interaction between surface-tethered 75-mer double strand DNA and integration host factor (IHF) protein was probed on Si-SiO2 substrates by determining the axial position of fluorescent labels attached to the free ends of DNA molecules.
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Despite its sub-nanometer precision axial localization capability, SSFM lacks high lateral resolution due to the low-NA requirement for planar reflectors. We developed a second technique, 4Pi-SSFM, which improves the lateral resolution of a conventional SSFM system by an order of magnitude while achieving nanometer-scale axial localization precision. Using two opposing high-NA objectives, fluorescence signal is interferometrically collected and spectral interference pattern is recorded. Axial position of emitters is found from analysis of the spectra. The 4Pi-SSFM technique was experimentally demonstrated by determining the surface profiles of fabricated glass surfaces and outer membranes of Shigella, a type of Gram-negative bacteria. A further discussion is presented to localize surface O antigen, which is an important oligosaccharide structure in the virulence mechanism of the Gram-negative bacteria, including E. coli and Shigella.
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