English
 
User Manual Privacy Policy Disclaimer Contact us
  Advanced SearchBrowse

Item

ITEM ACTIONSEXPORT
  Photobleaching and stabilization of fluorophores used for single-molecule analysis with one- and two-photon excitation

Dittrich, P. S., & Schwille, P. (2001). Photobleaching and stabilization of fluorophores used for single-molecule analysis with one- and two-photon excitation. Applied Physics B-Lasers and Optics, 73(8), 829-837. Retrieved from http://springerlink.metapress.com/content/tujupev7vfegk57d/fulltext.pdf.

Item is

Basic

show hide
Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0012-F50D-F Version Permalink: http://hdl.handle.net/11858/00-001M-0000-0027-E34C-8
Genre: Journal Article

Files

show Files
hide Files
:
16925.pdf (Publisher version), 0B
 
File Permalink:
-
Name:
16925.pdf
Description:
-
Visibility:
Restricted (Max Planck Institute for Biophysical Chemistry (Karl Friedrich Bonhoeffer Institute), Göttingen; )
MIME-Type / Checksum:
application/pdf
Technical Metadata:
Copyright Date:
-
Copyright Info:
-
License:
-

Locators

show

Creators

show
hide
 Creators:
Dittrich, P. S.1, Author              
Schwille, P.1, Author              
Affiliations:
1Research Group of Experimental Biophysics, MPI for biophysical chemistry, Max Planck Society, ou_578554              

Content

show
hide
Free keywords: -
 Abstract: Fluorescence spectroscopic measurements at the single-molecule level usually require large absorption cross sections and fluorescence quantum yields for the dyes under study. In addition to these parameters, the collectable number of fluorescence photons and, thus, the signal-to-noise ratio of the measurement, is influenced by processes like triplet-state population and photobleaching, shifting the saturation threshold of the dye to lower excitation intensities. Confocal fluorescence correlation spectroscopy (FCS) is a versatile method to precisely determine photon emission rates of single molecules but also gives access to rate constants of dynamic bleaching and intersystem crossing. In recent FCS studies in solution and living cells, two-photon excitation with its inherent spatial sectioning has proven to be a very valuable alternative to minimize background and cumulative signal loss. However, there is evidence that in many dye systems, the photobleaching rates with two-photon excitation are significantly enhanced with respect to one-photon excitation at comparable photon-emission yields. The reasons have so far remained mainly speculative. In the present study, potential photobleaching pathways are investigated by adding chemical stabilizers and by working at different oxygen concentrations. The results suggest that the population of triplet states does not appear to be responsible for the limited emission rate with two-photon excitation. Rather, photobleaching pathways via the formation of radicals seem to be plausible causes for the signal limitation. Favorable conditions are discussed to maximize the overall photon-collection yield in two-photon experiments.

Details

show
hide
Language(s): eng - English
 Dates: 2001-12
 Publication Status: Published in print
 Pages: -
 Publishing info: -
 Table of Contents: -
 Rev. Method: -
 Degree: -

Event

show

Legal Case

show

Project information

show

Source 1

show
hide
Title: Applied Physics B-Lasers and Optics
Source Genre: Journal
 Creator(s):
Affiliations:
Publ. Info: -
Pages: - Volume / Issue: 73 (8) Sequence Number: - Start / End Page: 829 - 837 Identifier: -