Re: Highly diffusible dye for filling neurons in vivo
Posted by
mmodel on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Highly-diffusible-dye-for-filling-neurons-in-vivo-tp7586162p7586163.html
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Hi Peter,
You can obtain cell morphology by adding the dye Acid Blue 9 to the medium at ~7 mg/ml. The sample is placed in a shallow chamber. The dye stays outside the cells and does no harm to them. Then you collect a transmission image at 630 nm, where the dye absorbance
is the strongest. Where the cells are thicker the absorbing layer is thinner, and the image brightness quantitatively reflects cell thickness. If you want we can talk more about it, and I can send you some papers.
Mike Model
From: Confocal Microscopy List <[hidden email]> on behalf of Peter Rupprecht <[hidden email]>
Sent: Wednesday, December 21, 2016 3:50 AM
To: [hidden email]
Subject: Highly diffusible dye for filling neurons in vivo
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Dear dye experts,
I'm currently patching very small neurons (soma 5-10 um in diameter, dendrites much smaller) in living tissue. In addition to electrophysiological recordings, I'd like to get the morphology without fixing
the tissue, so I add a dye (until recently mostly Alexa 594) to the pipette solution.
However, since the neuronal processes are very small, it is very often only the soma and not the small dendrites that get filled by the dye.
Switching from Alexa 594 to Alexa 488 improved things a bit (probably since 488 is smaller?
https://en.wikipedia.org/wiki/Alexa_Fluor).
But maybe somebody on the list has an idea of an even smaller, highly diffusible, non-toxic dye that I could use for this purpose. Plus, it should of course work with regular 2P excitation (800-950 nm).
Any ideas?
Best,
Peter