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antibody purification

Frank O. Fackelmayer Frank.Fackelmayer at uni-konstanz.de
Tue Jan 25 09:44:09 EST 2000


Dave Parcej wrote:

> Alternatively, affinity purify your antibody. the easiest way is to adsorb
> your protein onto PVDF and after blocking (eg with dried milk), incubate
> with the serum. You can easily elute with 0.2M glycine. HCl pH2.5 but
> remember to neutralise with 1M tris immediately!! This works well and
> normally reduces the overall background compared to crude serum
> Dave
>

Unfortunately that does not work very well in removing anti-E.coli
antibodies, because the protein preparation used for purifying the antibody
is usually also from recombinant origin and therefore contains bacterial
proteins. Of course, making affinity purified antibodies is always a good
idea for improving results. In some cases, it may however be helpful to
include a immobilized E.coli lysate column as an additional purification
step.

Frank








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