*The excitation is much broader so that many dyes are excited at the same time. This is an advantag... background.
*The molecular brightness of the dyes is generally around one order of magnitude smalle... cule applications.
* The photobleaching of the dyes is much more pronounced. This again limits the nu... of fluorescence photons you can achieve from your dyes. Bleaching only takes place in the excitation vol
nstant is too similar to distinguish between both dyes in a mixture by means of standard FCS.\\
Nevertheless, the autocorrelation curves of both dyes differ in the microsecond range. ATTO655 shows pr... somerization.\\
The fluorescence lifetime of both dyes differ significantly (~1.8 ns for ATTO655 and ~1 ... s to define these filters:
* If the different dyes have been measured separately, and TCPSC curves o
* **Note** that the diffusion coefficients of all dyes are temperature dependent.
* FCS measurements a... e through the confocal volume. Examples for these dyes are
ATTO655 or ATTO425. Also dyes with a moderate triplet contribution can be
fitted with this model if... cell membrane is observed. This model is made for dyes moving within a plane,
e.g. modified lipids in a
ions between two molecules labeled with different dyes. Usually, the dyes attached to the two molecules differ in their absorption and emission properties but... abeled with both fluorophores.\\
FCCS calibration dyes are available e.g. from IBA ([[http://www.iba-lif... ally, a positive control is needed, in which both dyes are moving in a coordinated fashion.\\
Additional
ions between two molecules labeled with different dyes. Usually, the dyes attached to the two molecules of interest differ in their absorption and emission pr... abeled with both fluorophores.\\
FCCS calibration dyes are available e.g. from IBA ([[http://www.iba-lif
Some origins of multiexponetial decays for single dyes|Some origins of multiexponetial decays for pure dyes]]
===== Microscopy =====
[[http://www.picoquant
= Some origins of multiexponetial decays for pure dyes ======
The fluorescence lifetime of a dye measu... c angle in a perfectly aligned spectrometer pure dyes in homogeneous media may exhibit a multiexponenti
/7353/appnote_diffusioncoefficients.pdf| standard dyes for FCS]] and perform a series of measurements wh... Diffusion Coefficients of Organosoluble Reference Dyes in Organic Media Measured by Dual-Focus Fluoresce
/10.1088/0957-0233/27/2/027001
Pyridinium Styryl Dyes have also been used as IRF standard:
Lei Li, Men... lph Jimenez, Jianhua Xu\\
Using Pyridinium Styryl Dyes as the Standards of Time-Resolved Instrument Resp
c_fitting_script_Image_17.png?600 }}
* If both dyes can be fitted with the same model, click "Fit All... le, it is of course meaningless, as two different dyes were fitted.
* Place the mouse over the graph,