Evicting hitchhiker antigens from purified antibodies

Luhrs, Keith A.; Harris, Debra A.; Summers, Scott; Parseghian, Missag H.

doi:10.1016/j.jchromb.2009.03.042

Cited by 28 publications

(38 citation statements)

References 18 publications

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“…Contaminant-IgG association under physiological conditions is routine with anti-chromatin antibodies because chromatin structure is so highly conserved across phyla. Monoclonal antibodies specific for chromatin from one species bind host chromatin expelled from dead host cells during cell culture production [21][22][23]. Lacking specialized purification procedures to dissociate the wrong-species antigen from the antibodies, they cause immunopotency for their authentic-species antigen to vary from lot to lot by up to a factor of 12 [23].…”

Section: Amplification Of Host Contamination By Igg-chromatin Interacmentioning

confidence: 99%

Non-immunospecific association of immunoglobulin G with chromatin during elution from protein A inflates host contamination, aggregate content, and antibody loss

Gagnon

Nian

Yang

et al. 2015

Journal of Chromatography A

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Monoclonal IgG at pH 3.5 expressed a tendency to self-associate and associate non-specifically with surfaces, including the surfaces of precipitated chromatin heteroaggregates. The tendency was elevated with protein A-eluted IgG still in elution buffer (100mM acetate, pH 3.5). Association of IgG with chromatin elements under protein A elution conditions amplified host protein contamination of the elution fraction about 15-fold, caused formation of aggregates that persisted after pH neutralization, and imposed an approximate 5% loss on IgG recovery. Neutralization released eluted IgG from its low pH associations with chromatin and caused heteroaggregate remnants to associate into large particles easily removed by microfiltration. Most effective host contaminant clearance was achieved by filtration after neutralization to pH 5.5. All chromatin-mediated liabilities were suspended by extraction of chromatin heteroaggregates in advance of protein A.

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Section: Amplification Of Host Contamination By Igg-chromatin Interacmentioning

confidence: 99%

Non-immunospecific association of immunoglobulin G with chromatin during elution from protein A inflates host contamination, aggregate content, and antibody loss

Gagnon

Nian

Yang

et al. 2015

Journal of Chromatography A

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“…Recent publications on protein A affinity chromatography have revealed that most of the contaminants that co-elute with IgG do so because they form stable complexes with the product during cell culture production (29,30). DNA liberated from lysed cells acts as a liquid phase cation exchanger (CX).…”

Section: Size Exclusion Chromatography (Sec)mentioning

confidence: 99%

“…Product complexation with other contaminants may also occur via hydrophobic interactions, hydrogen bonding, or metal affinity interactions. These complexes can persist throughout the course of an entire purification process, imposing an artificial ceiling on the level of final product purity that can be obtained (29,30). There is no reason to presume that virus particles should be immune from this phenomenon.…”

Section: Size Exclusion Chromatography (Sec)mentioning

confidence: 99%

“…Both also weaken hydrophobic interactions, as do protein-stabilizing organic additives such as propylene glycol or low molecular weight polyethylene glycol, such as PEG-400, or those smaller. Use of decomplexant washes with bioaffinity has been observed to improve host cell protein clearance by a factor of 10 or more (29,30), but this is less important than the fact that the contaminants they displace may otherwise persist throughout an entire purification process. SEC does not support the application of washes per se, but there are two variations that support the same benefits without sacrificing buffer exchange into whatever formulation is desired.…”

Section: Size Exclusion Chromatography (Sec)mentioning

confidence: 99%

“…As noted in the discussion of SEC, stable complexes can form between biological products and various contaminants during cell culture production. Secondary washes are able to dissociate such complexes and more effectively remove the contaminants from a product bound to an affinity support (29,30). Immunoaffinity and similarly strong interactions are sufficiently robust that they can tolerate potent combinations of decomplexants, such as 1.5 M sodium chloride, 2 M urea, and 10 mM EDTA, which collectively suspend nonspecific electrostatic complexes, weak hydrophobic interactions, hydrogen bonding, and metal affinity interactions.…”

Section: Biospecific Affinity Chromatographymentioning

confidence: 99%

See 2 more Smart Citations

Chromatographic Purification of Virus Particles

Gagnon¹

2009

Encyclopedia of Industrial Biotechnology

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The application of chromatography represents an increasing trend in the field of virus purification. This article focuses on the unique constraints imposed by virus particles and how they relate to the strengths and limitations of various chromatography media. Specific attention is given to the contributions of different types of physical supports, including porous particles, monoliths, and adsorptive membranes. Media characteristics are also discussed in the context of purification process control, reproducibility, and process economics. Protocols are offered for conducting method development with size exclusion, ion exchange, hydrophobic interactions, hydroxyapatite, and bioaffinity chromatography, and guidelines are summarized for combining individual methods into integrated multistep purification procedures suitable for preparation of clinical material.

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Identification and characterization of host cell protein product‐associated impurities in monoclonal antibody bioprocessing

Levy

Valente

Choe

et al. 2013

Biotech & Bioengineering

152

201

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Downstream processing of monoclonal antibodies (mAbs) has evolved to allow the specific process for a new product to be developed largely by empirical specialization of a platform process that enables removal of impurities of different kinds. A more complete characterization of impurities and the product itself would provide insights into the rational design of efficient downstream processes. This work identifies and characterizes host cell protein (HCP) product associated impurities, i.e., HCP species carried through the downstream processes via direct interactions with the mAb. Interactions between HCP and mAbs are characterized using cross interaction chromatography under solution conditions typical of those used in downstream processing. The interacting species are then identified by two dimensional gel electrophoresis and mass spectrometry. This methodology has been applied to identify product associated impurities in one particular purification step, namely protein A affinity chromatography, for four therapeutic mAbs as well as the Fab and Fc domains of one of these mAbs. The results show both the differences in HCP-mAb interactions among different mAbs, and the relative importance of product association compared to co-elution in protein A affinity chromatography.

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Evicting hitchhiker antigens from purified antibodies

Cited by 28 publications

References 18 publications

Non-immunospecific association of immunoglobulin G with chromatin during elution from protein A inflates host contamination, aggregate content, and antibody loss

Non-immunospecific association of immunoglobulin G with chromatin during elution from protein A inflates host contamination, aggregate content, and antibody loss

Chromatographic Purification of Virus Particles

Identification and characterization of host cell protein product‐associated impurities in monoclonal antibody bioprocessing

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