eGFP is detected in red channel confocal

[Wei]

Hello, I would like to know if anyone have similar experiences to share? In eGFP and Tdtomato double transgenic zebrafish larvae (12 dpf from two different promoters from different genes), eGFP was always detected weakly in the red channel but not vice versa. It looks like eGFP expressing cells are also expressing Tdtomato weakly. Tried to do ICC by double antibody staining but could not find good antibody against Tdtomato. Please share with me too if you have good experience with Tdtomato antibody (manufacturer or catalog#?).

Thank you so much for your sharing!

Lucas

[Sahar Tavakoli]

Hi Lucas,

It could be noise, as weak signals and the other FP are co-localized. Do you use the separate filter for each channel. E. g when you are imaging for GFP change the excitation filter for GFP excitation wavelength. So, any other wavelengths except GFP would be omitted. And the same for dTomate.

Good luck

Sahar

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From: Wei <iloveque...@gmail.com>
To: Zbra...@magpie.bio.indiana.edu
Sent: Friday, May 18, 2012 11:41 AM
Subject: [Zbrafish] eGFP is detected in red channel confocal

Hello, I would like to know if anyone have similar experiences to share? In eGFP and Tdtomato double transgenic zebrafish larvae (12 dpf from two different promoters from different genes), eGFP was always detected weakly in the red channel but not vice versa. It looks like eGFP expressing cells are also expressing Tdtomato weakly. Tried to do ICC by double antibody staining but could not find good antibody against Tdtomato. Please share with me too if you have good experience with Tdtomato antibody (manufacturer or catalog#?).

Thank you so much for your sharing!

Lucas

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