Production of Antibody Fab′ Fragments in<i>E. coli</i>

Humphreys, David P.; Bowering, Leigh C.

doi:10.1002/9780470485408.ch27

Cited by 4 publications

(5 citation statements)

References 214 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Although the majority of this Fab was present in the periplasmic (OS1) fraction, the yield was too low to be commercially viable (a maximum of 1 mg L −1 ), especially when compared with other expression data in the literature (e.g. Humphreys and Bowering). In addition, the quantity of Fab D1.3 in the culture broth rapidly increased towards the end of the fermentation, signifying cell lysis.…”

Section: Resultsmentioning

confidence: 99%

“…Complete cell lysis by mechanical (e.g. high pressure homogenization) or non‐mechanical means, required for release of cytoplasmic proteins, is not needed; rather, the outer membrane can be stripped away using osmotic shock or mild heat treatments …”

Section: Introductionmentioning

confidence: 99%

“…high pressure homogenization) or non-mechanical means, 7 required for release of cytoplasmic proteins, is not needed; rather, the outer membrane can be stripped away using osmotic shock 8 or mild heat treatments. 9,10 The resultant periplasmic extract is thereby already enriched for the recombinant protein of interest, the periplasm containing only around 4% to 8% of the natural E. coli cellular proteins, making purification simpler. 11 The periplasmic extract is also of lower volume than a whole cell lysate.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Periplasmic expression in and release of Fab fragments from Escherichia coli using stress minimization

Hsu

Thomas

Overton

2015

J of Chemical Tech & Biotech

View full text Add to dashboard Cite

BACKGROUND: The bacterium Escherichia coli is a commonly used host for the production of recombinant protein biopharmaceutical products. One class of such molecules is antibody fragments, typified by the Crohn's disease and rheumatoid arthritis therapy Certolizumab pegol (Cimzia ® ). Antibody fragments generated in E. coli are often directed to the periplasm, so that disulphide bonding can occur and release can be simplified. However, many recombinant protein products are prone to misfolding and mislocalization. Here, we optimized the production of a Fab fragment, D1.3, and its release from the periplasm of E. coli using osmotic shock.

show abstract

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Periplasmic expression in and release of Fab fragments from Escherichia coli using stress minimization

Hsu

Thomas

Overton

2015

J of Chemical Tech & Biotech

View full text Add to dashboard Cite

show abstract

“…Rational design of expression constructs is generally successful for targets with a well-known and conserved domain organization, such as antibodies (Humphreys and Bowering, 2009) or simple enzymes (Au et al, 2006). In contrast, the approach tends to fare less well for proteins that are unrelated to known structures or those with a poorly-understood domain organization, such as viral polyproteins or complex enzymes from higher organisms (Littler, 2010).…”

Section: Engineering Proteins For Structural Studies: Construct Desigmentioning

confidence: 98%

Engineering human MEK-1 for structural studies: A case study of combinatorial domain hunting

Meier

Brookings

Ceska

et al. 2012

Journal of Structural Biology

View full text Add to dashboard Cite

“…The design of the purification process is influenced by the physicochemical properties of the product and the nature of the process-and product-related impurities [9]. Despite small variations in the chemical nature of mAb, many companies have defined platform purification processes [10][11][12].…”

Section: Design Of Generic Process Platformsmentioning

confidence: 99%

Equipment design and facility layout for flexible biomanufacturing processes

Peuker¹,

Bogner²

2011

Engineering in Life Sciences

View full text Add to dashboard Cite

In process development and manufacturing, the biopharmaceutical industry requires high flexibility in its production facilities. These suites must be capable of producing clinical material or even drug substances for the market. Several products are manufactured in parallel or shortly after one another, which is challenging especially for the equipment. Product titers are increasing due to improved cell line performances, and personalized medicine will lead to tailor‐made drugs for a smaller group of patients. The resulting reduced upstream volumes enhance the opportunities for complete single‐use manufacturing trains. These requirements must be considered already during the design phase of the manufacturing facility. Engineering efforts have to be reduced as much as possible in order to reduce cost and timelines. Generic platform concepts for the overall process enable a shorter execution time as well as more efficient qualification procedures. In this contribution, the focus is on production processes for monoclonal antibodies (mAb) as a major product class of the biopharmaceutical industry. Two case studies introduce process platform concepts integrating single‐use equipment for an existing building and for a greenfield facility.

show abstract

Production of Antibody Fab′ Fragments inE. coli

Cited by 4 publications

References 214 publications

Periplasmic expression in and release of Fab fragments from Escherichia coli using stress minimization

Periplasmic expression in and release of Fab fragments from Escherichia coli using stress minimization

Engineering human MEK-1 for structural studies: A case study of combinatorial domain hunting

Equipment design and facility layout for flexible biomanufacturing processes

Contact Info

Product

Resources

About