You may wish to refer to several of the excellent review articles on
determination of host cell proteins in recombinant products.
A good place to start is
L. C. Eaton, "Host Cell Protein Assay Development for Recombinant
Pharmaceuticals," J. Chromatography, A, Vol 705(1), p. 105 (1995).
As discussed in this review, there really is no such thing as a "generic"
anti-E.coli protein antibody for the purpose you describe. Antibody pools
raised against E. coli are against the most antigenic cell proteins, and
more weakly antigenic proteins are poorly represented in the mix.
To further complicate matters, any purification process selects for some
host cell proteins and against others. One doesn't know whether the
contaminants appearing in the product have a stoichiometric amount of
antibody present in the mix. Obviously, absence/shortage of antibody to
any specific impurity leads to underestimation of that component.
To get around these problems, a cascaded series of immunizations can be
done to generate a population which contains antibodies against all the
detectable antigens (i.e., detectable by silver staining on 2D gels),
including the weakly antigenic ones. This type of work is described fully
in the reference cited above.
To simplify this effort, the immunization work is generally done by using
a subset of host cell proteins. Often, a null strain (E. coli of the same
strain as used in the production construct, but without the plasmid) is
grown up and the resultant proteins are fractionated by one or more steps
of the process used to purify the recombinant product. Thus, the pool of
reagent antibodies generated for each product is highly strain- and
process-specific.
Hope this helps.
John Hoogerheide
Analytical Methods and Services
Pharmacia & Upjohn
______________________________ Reply Separator _________________________________
Subject: Residual host cell protein
Author: Ed Cheung <ECheung@biomira.com> at INTERNET
Date: 1/14/1998 2:30 PM
We are interested in measuring residual host cell protein as trace
impurity in a recombinant product. Does anyone know of a contract lab
that offer this type of analysis ? Or is there a commercially available
anti-E.coli-protein antibody for a simple EIA test for the presence of
host cell protein ?
Thanks in advance.
Ed Cheung, Ph.D.
Analytical Development,
echeung@biomira.com
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