Halo/SNAPtag vs GFP

[simon walker (BI)-2]

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Dear List,
Advice please.  We're at the planning stage of some exps where we're going
to try and image fluorescently-labelled proteins expressed at low level in live
cells from a knock-in mouse.  We're going to need the signal to be as bright as
possible and we're weighing up the pros and cons of Halo/SNAPtag vs
fluorescent protein labelling.  Clearly there are advantages in the flexibility of
labels that Halo/SNAPtag gives, but in terms of absolute signal is there a clear
winner here?  I'm sure there are good theoretical arguments based on
quantum yield, stoichiometry etc as to why one should be better, but has
anyone directly compared the fluorescence signal from a Halo/SNAPtag
labelled protein against a fluorescent protein label?
Thanks,
Simon

[Fassler, Matthias]

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Hi Simon,

I have been recently working with the HaloTag although I did no direct comparison to any FP. I mainly used the HaloTag-TAMRA ligand and to a lesser extend the HaloTag-AF488 ligand in combination with slightly overexpressed proteins. Based on older experiments with GFP, my subjective impression is that the Halo Tag might be slightly brighter. However, I think the main advantage is really the flexibility and the ability to selectively stain surface proteins, e.g. to study receptor internalization.

Best regard
Matthias

Dr. Matthias Faßler | Application Scientist
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-----Original Message-----
From: Simon Walker [mailto:[hidden email]]
Sent: Wednesday, August 01, 2012 11:13 AM
Subject: Halo/SNAPtag vs GFP

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
*****

Dear List,
Advice please.  We're at the planning stage of some exps where we're going to try and image fluorescently-labelled proteins expressed at low level in live cells from a knock-in mouse.  We're going to need the signal to be as bright as possible and we're weighing up the pros and cons of Halo/SNAPtag vs fluorescent protein labelling.  Clearly there are advantages in the flexibility of labels that Halo/SNAPtag gives, but in terms of absolute signal is there a clear winner here?  I'm sure there are good theoretical arguments based on quantum yield, stoichiometry etc as to why one should be better, but has anyone directly compared the fluorescence signal from a Halo/SNAPtag labelled protein against a fluorescent protein label?
Thanks,
Simon

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