Application of nanoapertures to fluorescence fluctuation correlation spectroscopy
Abstract/Contents
- Abstract
- Fluorescence Fluctuation Correlation Spectroscopy (FCS) is a single molecule spectroscopic technique that uses the correlations in photon arrival time to compute the diffusion coefficient and concentration of diffusing dye molecules. The technique is handicapped by the diffraction limit in traditional confocal optical microscopes because low concentrations of about pico-molar to nano-molar levels are required to maintain single molecule levels. Thus, biologically relevant concentrations at micro-molar and above are not accessible without some method of reducing the observation volume. A method using nano-apertures to confine the excitation and emission volume is proposed to solve the low concentration requirement in FCS. Near-field electromagnetic field enhancements in c-shaped apertures will be exploited to overcome the signal fall off as the size of the apertures is decreased as higher concentration levels are desired, while maintain single molecule levels inside the observation volume. Experiments using square and c-shaped apertures milled in metallic thin films show strong wavelength dependence and initial experiments in spin coated R6G shows about 10 fold enhancement in the overall signal in c-shaped apertures over square shaped apertures of the same equivalent cross-sectional area. Further, an FCS experiment was carried out using Atto655 dye to confirm the focal volume reduction and fluorescence enhancement. Photon counting histogram techniques were used to reveal the detailed mechanism of the aperture resonance and the fluorescence enhancement. Finally, an application to pulsed interleaved excitation Forster resonance energy transfer (PIE-FRET) in ssDNA (single stranded DNA) pairs was carried out at a high enough concentration to stabilize the hybridized dsDNA (double stranded DNA). The nano-aperture results show that the PIE-FRET technique can be applied to solutions containing FRET pairs at 1000x the concentration than previously possible using diffraction limited focal volumes.
Description
| Type of resource | text |
|---|---|
| Form | electronic; electronic resource; remote |
| Extent | 1 online resource. |
| Publication date | 2014 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Associated with | Yuen, Max |
|---|---|
| Associated with | Stanford University, Department of Applied Physics. |
| Primary advisor | Hesselink, Lambertus |
| Thesis advisor | Hesselink, Lambertus |
| Thesis advisor | Brongersma, Mark L |
| Thesis advisor | Byer, R. L. (Robert L.), 1942- |
| Advisor | Brongersma, Mark L |
| Advisor | Byer, R. L. (Robert L.), 1942- |
Subjects
| Genre | Theses |
|---|
Bibliographic information
| Statement of responsibility | Max Yuen. |
|---|---|
| Note | Submitted to the Department of Applied Physics. |
| Thesis | Thesis (Ph.D.)--Stanford University, 2014. |
| Location | electronic resource |
Access conditions
- Copyright
- © 2014 by Max Yuen
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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