ER tracker Dyes

Model Organism: Worms Methods & Reagents
1

Hello everybody,

I am working with a ER resident protein. We have the trangenic with a GFP fusion and I would like to have another marker to confirm the localization.
In our lab we have the “ER-Tracker TMRed (glibenclamide BODIPY TR)”. Does anybody know a protocol to use it with C. elegans?

Does anybody know a protocol for ER staining in the worm?

Many thanks in advance,

Laura

2

I can’t believe you have had no replies to this post, given the number of researchers in the worm community swarming over the ER…

(that first line should hook a few from Wormbase)

I’ve never used ER-Tracker TMRed (glibenclamide BODIPY TR), so much of what I am about to say is born more out my curiosity than providing you with a definitive answer.

It’s used primarily as a live-cell stain isn’t it, although, with some caveats you can also pre-fix. So, you have two choices really;

  1. use one of the literally 100s of protocols that exist out there for fixing and staining worms and then follow the Molecular Probes protocol from a suitable point.

  2. try staining live worms with the Tracker dye in a small amount of S buffer (wow this Tracker stuff is expensive)…with the hope that the dye gets transported and targeted to the ER.

Just out of interest, doesn’t glibenclamide screw up certain forms of K+ channels and therefore possibly might that not alter ER function? Just a thought…

Steve

3

I agree with Steve’s suggestions, you could also try using a genotype that has a pretty permeable cuticle to facilitate easy/fast/robust uptake of dyes and drugs like e2698

4

Many thanks for both replies.

Finally the way it worked was fixing worms first. I haven’t optimized the concentration and incubation time yet, too much red for now.
I will let you know the conditions if you are interested in!

Kind regards,

Laura

5

Hey Laura,

I’m sure many members of the community (and future members!) would be grateful if you shared your protocol on the forum.

Thanks!