Cytiva eBook - 4

Better coverage assays for better HCP ELISAs: Quantitating the benefits
Measuring methods
At present, ELISAs (enzyme-linked immunosorbent assays) represent the gold standard for industrial HCP quantification. The concept is straightforward. Researchers
gather proteins from the unmodified cell line and raise a population of polyclonal
antibodies against them in an animal host. These antibodies are used to create an
HCP ELISA able to bind and quantify the HCPs present in culture fluid and inprocess
samples. Process developers then use the ELISA to measure the HCP levels to check
how effectively the purification process is removing the HCPs.
The approach is widely used in industry. Philip Ridley-Smith, Cobra Biologics, tells
GEN "that his company uses an ELISA method for the measurement of residual
Escherichia coli HCP in purified plasmid DNA bulk."
The popularity of the approach is also noted by James Graham, D Phil, technical director,
analytical R&D, at Lonza Pharma & Biotech. "ELISA is the current industry standard for release
testing of manufacturing batches," he says, "as described in U.S. Pharmacopeia Chapter 1132."
Other methods, such as liquid chromatography-mass spectrometry (LC-MS), have
the potential to be used to analyze harvested cell culture fluid (HCCF) and provide
quantitative information on HCPs.
These methods, however, have drawbacks. "Mass spectrometry," says Anne-Sophie
Brès, Global Product Manager at Cytiva, "requires a highly skilled operator and access
to expensive equipment. At present, the process is too time-consuming and costly
to be practical."
Assay challenges
Despite representing the gold standard, HCP ELISAs have limitations. No assay can
detect every HCP made by a cell line because of the way its antibodies are produced.
ELISA antibodies are made by injecting animals with HCPs extracted from a cell line.
The animal responds by producing antibodies that bind foreign proteins, enabling
their removal by the immune system. These antibodies form the basis of the assay.
HCP ELISAs do not have antibodies to all HCPs because immunized animals do not
mount an equal immune response to every single protein. Put simply, some HCPs
are more immunogenic than others.
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Amersham™ HCPQuant CHO host cell protein ELISA kit coated in 96-well strip plate.



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Cytiva eBook

Table of Contents for the Digital Edition of Cytiva eBook

Contents
Cytiva eBook - 1
Cytiva eBook - 2
Cytiva eBook - 3
Cytiva eBook - Contents
Cytiva eBook - 5
Cytiva eBook - 6
Cytiva eBook - 7
Cytiva eBook - 8
Cytiva eBook - 9
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Cytiva eBook - 11
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Cytiva eBook - 18
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