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Why, why, why...ELISA? A look at the benchmark HCP assay
Process-specific assays are the most specialized type.
Each is built around a specific product. Often developed by specialist providers, these assays account for
the upstream and downstream processes specific to a
product. In theory, the process-specific approach could
provide the most complete quantitation of HCPs possible by ELISA for that product, but it requires a considerable time and cost investment.
Platform assays can be developed in-house to suit a
specific expression platform, such as a variant of a cell
line, reducing the need for generic and process-specific
assays. The strength of this approach is in broad coverage and high sensitivity, attributes that make platform
assays applicable to multiple products manufactured
using the same cell line and similar processes.
Once platform assays are developed, U.S. Pharmacopeia
Chapter 1132 suggests they can be validated for subsequent products, and they can take the place of generic
assays through all stages of development, reducing the
per-product investment.
Not-so-complete coverage?
Despite being benchmark assays, ELISA can't detect
everything. It's unlikely you would get 100% coverage
of all HCPs using the raised antibodies, even with the
process-specific approach.

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Some HCPs won't be immunogenic enough to trigger
a response in the animal host, or will trigger only a low
response. Any antibodies with low affinity for the antigen might not be detectable by ELISA at all.
So, to use ELISAs for HCP quantitation, regulatory bodies ask that each assay be validated. In many protocols,
this involves running 2D-gel electrophoresis/Western
blot assays and comparing how well they detect
anti-HCP antibodies compared to an in-gel total
protein stain.
This type of coverage assay enables you to estimate a
percentage coverage, and it can direct you toward using a more specific ELISA or orthogonal assays, such as
mass spectrometry, that could fill the gap.
The increasing significance of checking coverage is
shown by updates to the U.S. Pharmacopeia in 2016
and the Pharmacopoeia Europaea in 2017 (USP 1132
and Ph. Eur. 2.06.34). A Japanese pharmacopeia update
on the same subject is under preparation.
Although these updates are just guidelines and best
practices, most analytical scientists now consider coverage assays an essential part of product development. n
LEARN MORE ABOUT Cytiva's
END-TO-END SOLUTION FOR HCP ANALYSIS

https://www.ncbi.nlm.nih.gov/pubmed/25998019 https://www.ncbi.nlm.nih.gov/pubmed/25998019 https://www.ncbi.nlm.nih.gov/pubmed/25998019 http://www.usp.org/biologics/proteins http://www.usp.org/biologics/proteins https://www.pmda.go.jp/rs-std-jp/standards-development/jp/pub-comments/jp/0147.html https://www.pmda.go.jp/rs-std-jp/standards-development/jp/pub-comments/jp/0147.html https://www.cytivalifesciences.com/solutions/protein-research/hcp-analysis?extcmp=CY20363-GL-GCR-PD-HCPprelaunchEbookGEN-exteml-exteml-ebook https://www.cytivalifesciences.com/solutions/protein-research/hcp-analysis?extcmp=CY20363-GL-GCR-PD-HCPprelaunchEbookGEN-exteml-exteml-ebook http://www.GENengnews.com

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Table of Contents for the Digital Edition of Cytiva eBook

Contents
Cytiva eBook - 1
Cytiva eBook - 2
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Cytiva eBook - Contents
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