Re: Fluorophore excitable with 405nm and 488nm lines

Posted by Martin Wessendorf-2 on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Fluorophore-excitable-with-405nm-and-488nm-lines-tp6432486p6432885.html

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Dear Daniel--

On 6/2/2011 1:59 PM, Daniel Gitler wrote:

> I am in need of either beads or a fluorophore in solution which is exited consistently by both the 405nm and 488nm lines of a confocal microscope. What I really need is that the ratio of excitation should be constant, in which case two separate dyes are probably not a good choice, unless their molar ratio can be quite consistent (perhaps in beads?). I also need that the efficiency of excitation for both lines be quite decent (doesn't have to be maximal, just decent). Finally (it appears I have a lot of requisites) the dye/beads should be as insensitive as possible to environmental changes (i.e., pH etc). The idea is to have a good standard in order to calibrate the ratio of the power of these two lines when imaging a sample (I do not need to know the actual number, just a relative measure will do fine).

Based on a quick search, uranium glass looks like a possibility.  The
absorbance is perhaps 2x better at 488 than 405.  However, if you're
imaging with a confocal, you would probably need to use point-scanning,
since uranium has a loooong fluorescence time-constant (e.g. 170 usec in
solution).

Here are a few (mediocre) references--they might be a starting point:

http://pubs.acs.org/doi/abs/10.1021/ac00230a029
http://webhost.bridgew.edu/cnoda/research/uranyl/index.html
http://onlinelibrary.wiley.com/doi/10.1002/pssa.2211150136/pdf

Good luck!

Martin
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