Posted by
Arne Seitz on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Several-Alexa-Fluor-dyes-extinction-coefficients-and-quantum-yields-tabulated-new-deconvolution-papes-tp7581205p7581237.html
*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy*****
Dear Gabor,
in the Handbook of Biological Confocal Microscopy in Chapter 16 ("Fluorophores for Confocal Microscopy: Photophysics and Photochemistry" by R. Tsien, L. Ernst, A. Waggoner) you can find a nice calculation of the efficiency of confocal systems. In essence: Per fluorophore you obtain photoelectron per sweep. This might serve a s a good starting point to make calculations for WF systems.
For the single molecule experiment you should be able to measure the photons which arrive at the camera (as done in many point localization papers). Knowing the fluorophore and the excitation intensity (at least roughly) you should be able to calculate the efficiency of the microscope pathway.
Have a nice day.
Cheers
Arne
> -----Original Message-----
> From: Confocal Microscopy List
> [mailto:
[hidden email]] On Behalf Of Csúcs
> Gábor
> Sent: jeudi 7 novembre 2013 13:34
> To:
[hidden email]
> Subject: Photon detection efficiency of microscopes
>
> *****
> To join, leave or search the confocal microscopy listserv, go to:
>
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy> *****
>
> Dear All,
>
> I made a quick search but found nothing really useful. My question is the
> following: does anyone have some data/literature how efficient microscopes
> are in detecting the photons of a fluorescent sample? Obviously the NA and
> the QE of the camera (plus the performance filters/dichroics) are important
> factors but taken e.g. A single molecule experiment with a 1.49 NA objective
> and a back-illuminated EM-CCD what ration of the emitted photons really
> reach the detector? Of course the transmission of the objectives and other
> lenses in the microscopes can vary but is there still an estimate?
>
> Thanks Gabor