fluorescein diacetate (fda)

[Aryeh Weiss]

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I am using fluorescein diacetate in my teaching lab. The student get to
see the cells light up, measure the increase over time (takes a minute
or two), see it photobleach afterwards, and  process it by subtracting
the in-cell signal from background fluorescence. Very nice.

My question concerns the storage of the fda. We made a 0.5mM stock
solution in acetone, which worked great with a further 1/200 dilution
into the plate. We could probably have also diluted 1/1000.
We stored the leftover stock solution at -20C. A week later that stock
solution did not work at all. When we added 10ul to cell culture plate ,
we saw an increase in background, but absolutely nothing in the cells.

We then made a fresh stock solution of fda, and it worked perfectly.
Exactly as expected.

So my question is - what happened?

Is there  a trick to keeping the stock solution active, or do we need to
make it fresh every week?

Thanks for any advise you can provide.

--aryeh

--
Aryeh Weiss
Faculty of Engineering
Bar Ilan University
Ramat Gan 52900 Israel

Ph:  972-3-5317638
FAX: [phone]

[Jacqueline Ross]

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Hi Aryeh,

For CMDFA, we make up the stock solution in dry DMSO as per the instruction sheet and then store in aliquots at -20 degrees Celsius. If it gets any water in it, it won’t work as it hydrolyses so we make sure to use freezer tubes that seal really tight, rather than standard Eppendorfs. It might be the same for FDA.

Cheers,

Jacqui

From: Confocal Microscopy List <[hidden email]> On Behalf Of Aryeh Weiss
Sent: Tuesday, April 20, 2021 9:33 PM
To: [hidden email]
Subject: fluorescein diacetate (fda)

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To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy<http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy>
Post images on http://www.imgur.com<http://www.imgur.com> and include the link in your posting.
*****

I am using fluorescein diacetate in my teaching lab. The student get to
see the cells light up, measure the increase over time (takes a minute
or two), see it photobleach afterwards, and  process it by subtracting
the in-cell signal from background fluorescence. Very nice.

My question concerns the storage of the fda. We made a 0.5mM stock
solution in acetone, which worked great with a further 1/200 dilution
into the plate. We could probably have also diluted 1/1000.
We stored the leftover stock solution at -20C. A week later that stock
solution did not work at all. When we added 10ul to cell culture plate ,
we saw an increase in background, but absolutely nothing in the cells.

We then made a fresh stock solution of fda, and it worked perfectly.
Exactly as expected.

So my question is - what happened?

Is there  a trick to keeping the stock solution active, or do we need to
make it fresh every week?

Thanks for any advise you can provide.

--aryeh

--
Aryeh Weiss
Faculty of Engineering
Bar Ilan University
Ramat Gan 52900 Israel

Ph: 972-3-5317638
FAX: [phone]