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Purification of recombinant protein A by aqueous two-phase extraction integrated with affinity precipitation (1992)

Kamihira, Masamichi ; Kaul, Rajni ; Mattiasson, Bo

New York, NY [u.a.] : Wiley-Blackwell

Biotechnology and Bioengineering 40 (1992), S. 1381-1387

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ISSN: 0006-3592

Keywords: aqueous two-phase extraction ; affinity precipitation ; Eudragit ; protein A ; Chemistry ; Biochemistry and Biotechnology

Source: Wiley InterScience Backfile Collection 1832-2000

Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology

Notes: Aqueous two-phase extraction incorporated affinity precipitation was examined as a technique for protein purification. An enteric coating polymer, Eudragit S100, was employed as a ligand carrier. Eudragit was specifically partitioned to the top phase in the aqueous two-phase systems. For application of this method to purification of recombinant protein A using human IgG coupled to Eudragit in an aqueous two-phase system, 80% of protein A added was recovered with 81% purity. The purity was enhanced 26-fold by thid method. The IgG-Eudragit could be used repeatedly for the purification process. This seperation method should be applicable to industrial-scale purification as a new purification procedure combining the advantages and compensating for the disadvantages of the aqueous two-phase method and affinity precipitation method. © 1992 John Wiley & Sons, Inc.

Additional Material: 5 Ill.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1002/bit.260401112

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Recovery of a monoclonal antibody from hybridoma culture supernatant by affinity precipitation with Eudragit S-100 (2000)

Ângela Taipa, M. ; Kaul, Rajni-Hatti ; Mattiasson, Bo ; [et al.]

Springer

Bioseparation 9 (2000), S. 291-298

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ISSN: 1573-8272

Keywords: affinity precipitation ; Eudragit S-100 ; monoclonal antibodies ; purification

Source: Springer Online Journal Archives 1860-2000

Topics: Process Engineering, Biotechnology, Nutrition Technology

Notes: Abstract An IgG1 monoclonal antibody (MAB) was isolated from hybridoma culture supernatant by affinity precipitation with an Eudragit S-100-based heterobifunctional ligand. Affinity binding was performed in a homogeneous aqueous phase at pH 7.5 followed by precipitation of the bound affinity complex by lowering the pH to 4.8. After two washing steps, elution of specifically bound MAB was achieved by incubating the precipitate with 0.1 M glycine.HCl pH 2.5. The influence of elution volume and time on the recovery of active MAB and the overall purification factor were studied. The best conditions enabled the recovery of 50.2% of active MAB with a purification factor of 6.2. A further dialysis against 50 mM Tris.HCl pH 8.0 increased the activity yield and the purification factor to 68.4% and 8.3, respectively. This result showed that part of the antibody activity loss during affinity precipitation was due to a reversible inactivation process, being easily recovered after a refining dialysis step.