Best Secondary Reagents for Rabbit Polyclonal Abs

Nebe-Von-Caron, G g.nebe-von-caron at unipath.com
Thu Jun 17 10:05:18 EST 2004


At last not a virus. 
I actually also use the Jackson donkey ant rabbit but in PE for surface
fluorescence with great success. Have previously used the sigma rabbit
anti-FITC monoclonal trying to follow some published protocol but that
was very poor in comparison. Have not tried the donkey FITC reagent.

Regards

Gerhard

-----Original Message-----
From: Hastings, Richard C [mailto:richard.hastings at astrazeneca.com] 
Sent: 16 June 2004 16:40
To: cyto-inbox
Subject: Best Secondary Reagents for Rabbit Polyclonal Abs


Hi,

Lately, I have been facsing many rabbit polyclonal antibodies. I have
been using Molecular Probes goat anti-rabbit IgG Alexa Fluor 488 and
Jackson Immunoresearch donkey F(ab')2 anti-rabbit IgG (H+L) FITC or APC
as secondary reagents. I use fluorochromes that don't overlap with
propidium iodide because I like using PI as a marker of cell death
(yeah, I know PI is more a marker of membrane permeability, so humor me
a bit here and I do know that APC and PI overlap in their emission
wavelengths). Problems I have run into are these secondary reagents are
not bright at all, the rabbit IgG preps tend to have a lot of
non-specific binding and purified rabbit IgG is a lousy control. 

I have also tried the Molecular Probes rabbit Zenon kit and the results
were less than satisfactory. Anyone have any suggestions?

Richard C. Hastings
Associate Scientist
Target Development/TBD
1800 Concord Pike
Wilmington, DE 19803
302-886-2452
richard.hastings at astrazeneca.com





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