| 1. |
- Blom, Hans, et al.
(författare)
-
Electrostatic Interactions of Fluorescent Molecules with Dielectric Interfaces Studied by Total Internal Reflection Fluorescence Correlation Spectroscopy
- 2010
-
Ingår i: International Journal of Molecular Sciences. - : MDPI. - 1661-6596 .- 1422-0067. ; 11:2, s. 368-406
-
Tidskriftsartikel (refereegranskat)abstract
- Electrostatic interactions between dielectric surfaces and different fluorophoresused in ultrasensitive fluorescence microscopy are investigated using objective-based TotalInternal Reflection Fluorescence Correlation Spectroscopy (TIR-FCS). The interfacialdynamics of cationic rhodamine 123 and rhodamine 6G, anionic/dianionic fluorescein,zwitterionic rhodamine 110 and neutral ATTO 488 are monitored at various ionic strengthsat physiological pH. As analyzed by means of the amplitude and time-evolution of theautocorrelation function, the fluorescent molecules experience electrostatic attraction orrepulsion at the glass surface depending on their charges. Influences of the electrostaticinteractions are also monitored through the triplet-state population and triplet relaxationtime, including the amount of detected fluorescence or the count-rate-per-moleculeparameter. These TIR-FCS results provide an increased understanding of how fluorophoresare influenced by the microenvironment of a glass surface, and show a promising approachfor characterizing electrostatic interactions at interfaces.
|
|
| 2. |
- Blom, Hans, et al.
(författare)
-
Triplet-State Investigations of Fluorescent Dyes at Dielectric Interfaces Using Total Internal Reflection Fluorescence Correlation Spectroscopy
- 2009
-
Ingår i: Journal of Physical Chemistry A. - Washington, DC : American Chemical Society. - 1089-5639 .- 1520-5215. ; 113:19, s. 5554-5566
-
Tidskriftsartikel (refereegranskat)abstract
- The triplet-state kinetics of several fluorescent dyes used in ultrasensitive fluorescence microscopy are investigated using total internal reflection fluorescence correlation spectroscopy (TIR-FCS). A theoretical outline of the correlation analysis and the physical aspects of evanescent excitation and fluorescence emission at dielectric interfaces are given. From this analysis, the rates of intersystem crossing and triplet decay are deduced for fluorescein, ATTO 488, rhodamine 110, rhodamine 123, and rhodamine 6G in aqueous buffer solutions. All investigated dyes show slightly higher triplet rates at the dielectric interface compared to bulk solution measurements. We attribute this enhancement to possible modifications of the dyes’ photophysical properties near a dielectric interface. In the case of rhodamine 6G, the impact of changes in the dye concentration, ionic strength of the solvent, and potassium iodide concentration are also investigated. This leads to a better understanding of the influences of dye−dye, dye−solvent, and dye−surface interactions on the increased triplet intersystem crossing and triplet decay rates. The study shows that analysis of triplet-state kinetics by TIR-FCS not only results in a better understanding of how the photophysical properties of the dyes are affected by the presence of an interface, but also provides a means for probing the microenvironment near dielectric interfaces.
|
|
| 3. |
- Hassler, Kai, et al.
(författare)
-
Dynamic disorder in horseradish peroxidase observed with total internal reflection fluorescence correlation spectroscopy
- 2007
-
Ingår i: Optics Express. - 1094-4087. ; 15:9, s. 5366-5375
-
Tidskriftsartikel (refereegranskat)abstract
- This paper discusses the application of objective-type total internal reflection fluorescence correlation spectroscopy (TIR-FCS) to the study of the kinetics of immobilized horseradish peroxidase on a single molecule level. Objective-type TIR-FCS combines the advantages of FCS with TIRF microscopy in a way that allows for simultaneous ultra-sensitive spectroscopic measurements using a single-point detector and convenient localization of single molecules on a surface by means of parallel imaging.
|
|
| 4. |
- Leutenegger, M., et al.
(författare)
-
Single molecule detection at surfaces : Dual-color fluorescence fluctuation spectroscopy with total internal reflection excitation
- 2007
-
Ingår i: New Approaches in Biomedical Spectroscopy. - Washington, DC : American Chemical Society (ACS). - 0841274371 - 9780841274372 ; , s. 259-279
-
Konferensbidrag (refereegranskat)abstract
- Fluorescence fluctuation spectroscopy based on an evanescent field excitation scheme leads to an alternative concept for single molecule detection with interesting experimental features. This concept is described and analyzed regarding the total fluorescence process including excitation, fluorescence emission and collection close to a dielectric interface. The realized experimental scheme showed superior performance of this concept with substantially enhanced molecular brightness when compared to the classical confocal setup based on water immersion objectives. Selected applications in life sciences including dual-color surface fluorescence correlation spectroscopy, enzyme kinetics and receptor-ligand binding are underlining the interest of this experimental approach.
|
|
| 5. |
- Sonesson, Andreas, et al.
(författare)
-
Protein-surfactant interactions at hydrophobic interfaces studied with Total Internal Reflection Fluorescence Correlation Spectroscopy (TIR-FCS)
- 2008
-
Ingår i: Journal of Colloid and Interface Science. - : Elsevier BV. - 0021-9797 .- 1095-7103. ; 317:2, s. 449-457
-
Tidskriftsartikel (refereegranskat)abstract
- The aim of this work was to study the dynamics of proteins near solid surfaces in the presence or absence of competing surfactants by means of total internal reflection fluorescence correlation spectroscopy (TIR-FCS). Two different proteins were studied, bovine serum albumin (BSA) and Thermomyces lanuginosus lipase (TLL). A nonionic/anionic (C12E6/LAS) surfactant composition was used to mimic a detergent formulation and the surfaces used were C 18 terminated glass. It was found that with increasing surfactant concentrations the term in the autocorrelation function (ACF) representing surface binding decreased. This Suggested that the proteins were competed off the hydrophobic surface by the surfactant. When fitting the measured ACF to a model for surface kinetics, it was seen that with raised C12E6/LAS concentration, the Surface interaction rate increased for both proteins. Under these experimental conditions this meant that the time the protein was bound to the surface decreased. At 10 mu M C12E6/LAS the surface interaction was not visible for BSA, whereas it was still distinguishable in the ACF for TLL. This indicated that TLL had a higher affinity than BSA for the C 18 surface. The study showed that TIR-FCS provides a useful tool to quantify the surfactant effect on proteins adsorption.
|
|
