Integrated isolation of antibody fragments from microbial cell culture fluids using supermacroporous cryogels
(2004) In Journal of Chromatography A 1045(1-2). p.93-98- Abstract
- The present paper describes a chromatographic capture/purification step for the recovery of proteins directly from undiluted and unclarified cell culture broths using supermacroporous dimethylacrylamide (DMAA) cryogel. The interconnected character and the size (10-100 mum) of the pores of the adsorbent make it possible to process whole cell fermentation broths without blocking the column. Cu2+-iminodiacetic acid (IDA) DMAA cryogel has been used for the isolation and purification of excreted (His)(6)-tagged single chain (sc) Fv antibody fragments, (His)(6)-scFv, from E. coli cell culture. Bound protein was recovered with 0.2 M imidazole or with 20 mM EDTA and was practically cell-free. Chromatographic capture using Cu2+-IDA cryogel column... (More)
- The present paper describes a chromatographic capture/purification step for the recovery of proteins directly from undiluted and unclarified cell culture broths using supermacroporous dimethylacrylamide (DMAA) cryogel. The interconnected character and the size (10-100 mum) of the pores of the adsorbent make it possible to process whole cell fermentation broths without blocking the column. Cu2+-iminodiacetic acid (IDA) DMAA cryogel has been used for the isolation and purification of excreted (His)(6)-tagged single chain (sc) Fv antibody fragments, (His)(6)-scFv, from E. coli cell culture. Bound protein was recovered with 0.2 M imidazole or with 20 mM EDTA and was practically cell-free. Chromatographic capture using Cu2+-IDA cryogel column was performed at flow rates of 300 and 600 cm/h, respectively and resulted in 84-96% recovery of (His)(6)-scFv fragments with a purification factor of 13-15. The DMAA cryogel adsorbent is mechanically stable, can withstand harsh cleaning-in-place procedure and is relatively inexpensive. Chromatographic isolation of proteins using cryogels allows efficient removal of cells and can be operated at a flow rate as high as 600 cm/h. This novel technique has proven to be a scalable process, does not require special equipment and can be a good alternative to expanded bed adsorption and other integrated isolation techniques. (C) 2004 Published by Elsevier B.V. (Less)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/140600
- author
- Dainiak, Maria LU ; Kumar, Ashok LU ; Plieva, Fatima LU ; Galaev, Igor LU and Mattiasson, Bo LU
- organization
- publishing date
- 2004
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- Cryogels, Antibodies, Proteins, Dimethylacrylamide
- in
- Journal of Chromatography A
- volume
- 1045
- issue
- 1-2
- pages
- 93 - 98
- publisher
- Elsevier
- external identifiers
-
- wos:000223212000011
- pmid:15378883
- scopus:3843130508
- ISSN
- 0021-9673
- DOI
- 10.1016/j.chroma.2004.06.029
- language
- English
- LU publication?
- yes
- id
- af7e72c3-bdea-402e-a55e-9d0f39713ee2 (old id 140600)
- date added to LUP
- 2016-04-01 15:42:33
- date last changed
- 2023-08-15 13:30:56
@article{af7e72c3-bdea-402e-a55e-9d0f39713ee2, abstract = {{The present paper describes a chromatographic capture/purification step for the recovery of proteins directly from undiluted and unclarified cell culture broths using supermacroporous dimethylacrylamide (DMAA) cryogel. The interconnected character and the size (10-100 mum) of the pores of the adsorbent make it possible to process whole cell fermentation broths without blocking the column. Cu2+-iminodiacetic acid (IDA) DMAA cryogel has been used for the isolation and purification of excreted (His)(6)-tagged single chain (sc) Fv antibody fragments, (His)(6)-scFv, from E. coli cell culture. Bound protein was recovered with 0.2 M imidazole or with 20 mM EDTA and was practically cell-free. Chromatographic capture using Cu2+-IDA cryogel column was performed at flow rates of 300 and 600 cm/h, respectively and resulted in 84-96% recovery of (His)(6)-scFv fragments with a purification factor of 13-15. The DMAA cryogel adsorbent is mechanically stable, can withstand harsh cleaning-in-place procedure and is relatively inexpensive. Chromatographic isolation of proteins using cryogels allows efficient removal of cells and can be operated at a flow rate as high as 600 cm/h. This novel technique has proven to be a scalable process, does not require special equipment and can be a good alternative to expanded bed adsorption and other integrated isolation techniques. (C) 2004 Published by Elsevier B.V.}}, author = {{Dainiak, Maria and Kumar, Ashok and Plieva, Fatima and Galaev, Igor and Mattiasson, Bo}}, issn = {{0021-9673}}, keywords = {{Cryogels; Antibodies; Proteins; Dimethylacrylamide}}, language = {{eng}}, number = {{1-2}}, pages = {{93--98}}, publisher = {{Elsevier}}, series = {{Journal of Chromatography A}}, title = {{Integrated isolation of antibody fragments from microbial cell culture fluids using supermacroporous cryogels}}, url = {{http://dx.doi.org/10.1016/j.chroma.2004.06.029}}, doi = {{10.1016/j.chroma.2004.06.029}}, volume = {{1045}}, year = {{2004}}, }