Epitope tags

WS via methods%40net.bio.net (by novalidaddress from nurfuerspam.de)
Mon Jun 6 04:50:30 EST 2011


Dear Engelbert,

do you have by chance an idea, how your precipitation protocol for IgY
purification performs in comparison to immobilized 2-mercaptopyrinide
affinity chromatography (the matrix GE
- formerly Amersham / Pharmacia - sells for IgY purification?). These
tiny pre-packed columns (5ml) are nice, but if you start with one egg,
you already will need for runs, as they only bind 25mg IgY per run.
Scale up is expensive therefore. I thought of bulk purification in the
>1g IgY scale (means 10 eggs and up)

Another question: How to avoid trace contamination with avidin? Is
extensive washing of the intact yolks in order to get rid of all egg
white remains sufficient or may I expect some avidin being present in
the yolk, too?

Thanks for your help!

Wolfgang

> Isolation of IgY is very simple, the yolk is diluted 10-fold with PBS and a 35-120 mg/ml
> PEG-6000 cut is prepared. The crude preparation is dissolved in PBS
> again and then further purified by a 25-50% ammonium sulphate cut. The
> precipitate stores well suspended in 50% AS in the fridge.


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