Continuous Fc detection for protein A capture process control

Patil, Ujwal; Crum, Mary; Vu, Binh; Wasden, Katherine; Kourentzi, Katerina; Willson, Richard C.

doi:10.1016/j.bios.2020.112327

Cited by 9 publications

(6 citation statements)

References 45 publications

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“…Briefly, a purified gene fragment (30 ng/μL) was incubated with NdeI/XhoI digested pET28a (50 ng/μL) and 2X GA reaction mix at 50 °C for 2 h. The same procedure was repeated to produce the cyclic Z3 protein-expressing plasmid pET28a-Z3cyc. Separately, linear Z3 protein-expressing plasmid, pET28a-Z3lin was produced according to our previously reported study [ 52 ].…”

Section: Methodsmentioning

confidence: 99%

Expression and Characterization of Intein-Cyclized Trimer of Staphylococcus aureus Protein A Domain Z

Nandy

Maranholkar

Crum

et al. 2023

IJMS

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Staphylococcus aureus protein A (SpA) is an IgG Fc-binding virulence factor that is widely used in antibody purification and as a scaffold to develop affinity molecules. A cyclized SpA Z domain could offer exopeptidase resistance, reduced chromatographic ligand leaching after single-site endopeptidase cleavage, and enhanced IgG binding properties by preorganization, potentially reducing conformational entropy loss upon binding. In this work, a Z domain trimer (Z3) was cyclized using protein intein splicing. Interactions of cyclic and linear Z3 with human IgG1 were characterized by differential scanning fluorimetry (DSF), surface plasmon resonance (SPR), and isothermal titration calorimetry (ITC). DSF showed a 5 ℃ increase in IgG1 melting temperature when bound by each Z3 variant. SPR showed the dissociation constants of linear and cyclized Z3 with IgG1 to be 2.9 nM and 3.3 nM, respectively. ITC gave association enthalpies for linear and cyclic Z3 with IgG1 of −33.0 kcal/mol and −32.7 kcal/mol, and −T∆S of association 21.2 kcal/mol and 21.6 kcal/mol, respectively. The compact cyclic Z3 protein contains 2 functional binding sites and exhibits carboxypeptidase Y-resistance. The results suggest cyclization as a potential approach toward more stable SpA-based affinity ligands, and this analysis may advance our understanding of protein engineering for ligand and drug development.

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Section: Methodsmentioning

confidence: 99%

Expression and Characterization of Intein-Cyclized Trimer of Staphylococcus aureus Protein A Domain Z

Nandy

Maranholkar

Crum

et al. 2023

IJMS

Self Cite

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“…In addition, Raman spectroscopy has been used for controlling lactate concentration which has led to improvements in cell density, viability, and protein production in mammalian cell culture 42 as well as with controlling glucose levels in the bioreactor 43, 44 . Using Raman spectroscopy to predict different mAb isotypes and glycosylation species with low prediction errors and through a permeate stream (without sample removal) has also been reported 45, 46 . A recent study also successfully implemented Raman spectroscopy for fine‐tuning mAb galactosylation levels in real‐time by controlling lactic acid feeding rate 47 .…”

Section: Integrated Process Analytical Technology (Ipat)mentioning

confidence: 99%

“…43,44 Using Raman spectroscopy to predict different mAb isotypes and glycosylation species with low prediction errors and through a permeate stream (without sample removal) has also been reported. 45,46 A recent study also successfully implemented Raman spectroscopy for fine-tuning mAb galactosylation levels in real-time by controlling lactic acid feeding rate. 47 However, this technique has certain limitations, such as giving highly overlapping spectral signatures that require deconvolution and having poor sensitivity to lower concentration species (which makes it difficult to continually observe all desired products).…”

Section: Upstream Downstreammentioning

confidence: 99%

Recent advances in integrated process analytical techniques, modeling, and control strategies to enable continuous biomanufacturing of monoclonal antibodies

Chopda

Gyorgypal

et al. 2021

J of Chemical Tech & Biotech

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Continuous bioprocessing is significantly changing the biological drugs (or biologics) manufacturing landscape by potentially improving product quality, process stability, and overall profitability, as was similarly seen during the adoption of advanced manufacturing processes for small molecule drugs in the past decade. However, the implementation of continuous manufacturing for biological processes producing protein-based drug molecules, such as monoclonal antibodies (mAbs), is facing several new hurdles. The barriers to continuous bioprocessing can be overcome through improved process understanding via better predictive capabilities enabled by hybrid modeling that can also lead to robust process control. This review article summarizes the recent advances and ongoing obstacles faced during the use of advanced process analytical technologies (PAT), process modeling, and control strategies to enable continuous manufacturing of mAbs. In addition, this review also discusses the process strategies and future directions of advanced continuous manufacturing approaches that have been adapted by other industries and that could be implemented for mAbs production soon.

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“…The use of the same specific ligand in the affinity column and in the sensor allowed simultaneous in‐line, one‐pot regeneration of column and sensor [137]. A fluorescence‐based monitoring technology allowing real‐time monoclonal antibody monitoring and for detecting IgG in column breakthrough was developed [138]. For this purpose, the column effluent was continuously contacted with soluble fluorescein‐labeled Fc‐binding ligands to produce an on‐line‐detectable shift in either fluorescence polarization or intensity [138].…”

Section: Bioaffinity Chromatography Techniquesmentioning

confidence: 99%

Recent trends in sorbents for bioaffinity chromatography

Kip

Hamaloğlu

Demir

et al. 2021

J of Separation Science

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Isolation or enrichment of biological molecules from complex biological samples is mostly a prerequisite in proteomics, genomics, and glycomics. Different techniques have been used to advance the efficiency of the purification of biological molecules. Bioaffinity chromatography is one of the most powerful technique that plays an important role in the isolation of target biological molecules by the specific interactions with ligands that are immobilized on different support materials. This review examines the recent developments in bioaffinity chromatography particularly over the past 5 years in the literature. Also properties of supports, immobilization techniques, types of binding agents, and methods used in bioaffinity chromatography applications are summarized.

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Continuous Fc detection for protein A capture process control

Cited by 9 publications

References 45 publications

Expression and Characterization of Intein-Cyclized Trimer of Staphylococcus aureus Protein A Domain Z

Expression and Characterization of Intein-Cyclized Trimer of Staphylococcus aureus Protein A Domain Z

Recent advances in integrated process analytical techniques, modeling, and control strategies to enable continuous biomanufacturing of monoclonal antibodies

Recent trends in sorbents for bioaffinity chromatography

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