Efficient enrichment of high-producing recombinant Chinese hamster ovary cells for monoclonal antibody by flow cytometry
To screen a high-producing recombinant Chinese hamster ovary (CHO) cell from transfected cells is generally laborious and time-consuming. We developed an efficient enrichment strategy for high-producing cell screening using flow cytometry (FCM). A stable pool that had possibly shown a huge variety o...
Ausführliche Beschreibung
Gespeichert in:
| Autor*in: |
Okumura, Takeshi [verfasserIn] |
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| Format: |
Artikel |
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| Sprache: |
Englisch |
| Erschienen: |
2015 |
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| Rechteinformationen: |
Nutzungsrecht: Copyright © 2015 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved. © COPYRIGHT 2015 Elsevier B.V. |
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| Schlagwörter: |
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| Übergeordnetes Werk: |
Enthalten in: Journal of bioscience and bioengineering - Osaka, 1999, 120(2015), 3, Seite 340 |
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| Übergeordnetes Werk: |
volume:120 ; year:2015 ; number:3 ; pages:340 |
| Links: |
|---|
| DOI / URN: |
10.1016/j.jbiosc.2015.01.007 |
|---|
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| 520 | |a To screen a high-producing recombinant Chinese hamster ovary (CHO) cell from transfected cells is generally laborious and time-consuming. We developed an efficient enrichment strategy for high-producing cell screening using flow cytometry (FCM). A stable pool that had possibly shown a huge variety of monoclonal antibody (mAb) expression levels was prepared by transfection of an expression vector for mAb production to a CHO cell. To enrich high-producing cells derived from a stable pool stained with a fluorescent-labeled antibody that binds to mAb presented on the cell surface, we set the cell size and intracellular density gates based on forward scatter (FSC) and side scatter (SSC), and collected the brightest 5% of fluorescein isothiocyanate (FITC)-positive cells from each group by FCM. The final product concentration in a fed-batch culture of cells sorted without FSC and SSC gates was 1.2-1.3-times higher than that of unsorted cells, whereas that of cells gated by FSC and SSC was 3.4-4.7-fold higher than unsorted cells. Surprisingly, the fraction with the highest final product concentration indicated the smallest value of FSC and SSC, and the middle value of fluorescence intensity among all fractionated cells. Our results showed that our new screening strategy by FCM based on FSC and SSC gates could achieve an efficient enrichment of high-producing cells with the smallest value of FSC and SSC. | ||
| 540 | |a Nutzungsrecht: Copyright © 2015 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved. | ||
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| 650 | 4 | |a Fluorescein | |
| 650 | 4 | |a Monoclonal antibodies | |
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| 700 | 1 | |a Masuda, Kenji |4 oth | |
| 700 | 1 | |a Watanabe, Kazuhiko |4 oth | |
| 700 | 1 | |a Miyadai, Kenji |4 oth | |
| 700 | 1 | |a Nonaka, Koichi |4 oth | |
| 700 | 1 | |a Yabuta, Masayuki |4 oth | |
| 700 | 1 | |a Omasa, Takeshi |4 oth | |
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10.1016/j.jbiosc.2015.01.007 doi PQ20160617 (DE-627)OLC1965022901 (DE-599)GBVOLC1965022901 (PRQ)g1424-5e69a1a7daf2db8085c07129bcb9aa3f95076b26af9c58682edf158fdc510f0f0 (KEY)0039398320150000120000300340efficientenrichmentofhighproducingrecombinantchine DE-627 ger DE-627 rakwb eng 570 540 660 DNB 42.00 bkl 58.30 bkl 48.00 bkl Okumura, Takeshi verfasserin aut Efficient enrichment of high-producing recombinant Chinese hamster ovary cells for monoclonal antibody by flow cytometry 2015 Text txt rdacontent ohne Hilfsmittel zu benutzen n rdamedia Band nc rdacarrier To screen a high-producing recombinant Chinese hamster ovary (CHO) cell from transfected cells is generally laborious and time-consuming. We developed an efficient enrichment strategy for high-producing cell screening using flow cytometry (FCM). A stable pool that had possibly shown a huge variety of monoclonal antibody (mAb) expression levels was prepared by transfection of an expression vector for mAb production to a CHO cell. To enrich high-producing cells derived from a stable pool stained with a fluorescent-labeled antibody that binds to mAb presented on the cell surface, we set the cell size and intracellular density gates based on forward scatter (FSC) and side scatter (SSC), and collected the brightest 5% of fluorescein isothiocyanate (FITC)-positive cells from each group by FCM. The final product concentration in a fed-batch culture of cells sorted without FSC and SSC gates was 1.2-1.3-times higher than that of unsorted cells, whereas that of cells gated by FSC and SSC was 3.4-4.7-fold higher than unsorted cells. Surprisingly, the fraction with the highest final product concentration indicated the smallest value of FSC and SSC, and the middle value of fluorescence intensity among all fractionated cells. Our results showed that our new screening strategy by FCM based on FSC and SSC gates could achieve an efficient enrichment of high-producing cells with the smallest value of FSC and SSC. Nutzungsrecht: Copyright © 2015 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved. © COPYRIGHT 2015 Elsevier B.V. Fluorescein Monoclonal antibodies Ovarian cancer Masuda, Kenji oth Watanabe, Kazuhiko oth Miyadai, Kenji oth Nonaka, Koichi oth Yabuta, Masayuki oth Omasa, Takeshi oth Enthalten in Journal of bioscience and bioengineering Osaka, 1999 120(2015), 3, Seite 340 (DE-627)265549817 (DE-600)1465387-4 (DE-576)074842005 1389-1723 nnns volume:120 year:2015 number:3 pages:340 http://dx.doi.org/10.1016/j.jbiosc.2015.01.007 Volltext http://www.ncbi.nlm.nih.gov/pubmed/25683450 GBV_USEFLAG_A SYSFLAG_A GBV_OLC SSG-OLC-TEC SSG-OLC-CHE SSG-OLC-FOR GBV_ILN_70 42.00 AVZ 58.30 AVZ 48.00 AVZ AR 120 2015 3 340 |
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10.1016/j.jbiosc.2015.01.007 doi PQ20160617 (DE-627)OLC1965022901 (DE-599)GBVOLC1965022901 (PRQ)g1424-5e69a1a7daf2db8085c07129bcb9aa3f95076b26af9c58682edf158fdc510f0f0 (KEY)0039398320150000120000300340efficientenrichmentofhighproducingrecombinantchine DE-627 ger DE-627 rakwb eng 570 540 660 DNB 42.00 bkl 58.30 bkl 48.00 bkl Okumura, Takeshi verfasserin aut Efficient enrichment of high-producing recombinant Chinese hamster ovary cells for monoclonal antibody by flow cytometry 2015 Text txt rdacontent ohne Hilfsmittel zu benutzen n rdamedia Band nc rdacarrier To screen a high-producing recombinant Chinese hamster ovary (CHO) cell from transfected cells is generally laborious and time-consuming. We developed an efficient enrichment strategy for high-producing cell screening using flow cytometry (FCM). A stable pool that had possibly shown a huge variety of monoclonal antibody (mAb) expression levels was prepared by transfection of an expression vector for mAb production to a CHO cell. To enrich high-producing cells derived from a stable pool stained with a fluorescent-labeled antibody that binds to mAb presented on the cell surface, we set the cell size and intracellular density gates based on forward scatter (FSC) and side scatter (SSC), and collected the brightest 5% of fluorescein isothiocyanate (FITC)-positive cells from each group by FCM. The final product concentration in a fed-batch culture of cells sorted without FSC and SSC gates was 1.2-1.3-times higher than that of unsorted cells, whereas that of cells gated by FSC and SSC was 3.4-4.7-fold higher than unsorted cells. Surprisingly, the fraction with the highest final product concentration indicated the smallest value of FSC and SSC, and the middle value of fluorescence intensity among all fractionated cells. Our results showed that our new screening strategy by FCM based on FSC and SSC gates could achieve an efficient enrichment of high-producing cells with the smallest value of FSC and SSC. Nutzungsrecht: Copyright © 2015 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved. © COPYRIGHT 2015 Elsevier B.V. Fluorescein Monoclonal antibodies Ovarian cancer Masuda, Kenji oth Watanabe, Kazuhiko oth Miyadai, Kenji oth Nonaka, Koichi oth Yabuta, Masayuki oth Omasa, Takeshi oth Enthalten in Journal of bioscience and bioengineering Osaka, 1999 120(2015), 3, Seite 340 (DE-627)265549817 (DE-600)1465387-4 (DE-576)074842005 1389-1723 nnns volume:120 year:2015 number:3 pages:340 http://dx.doi.org/10.1016/j.jbiosc.2015.01.007 Volltext http://www.ncbi.nlm.nih.gov/pubmed/25683450 GBV_USEFLAG_A SYSFLAG_A GBV_OLC SSG-OLC-TEC SSG-OLC-CHE SSG-OLC-FOR GBV_ILN_70 42.00 AVZ 58.30 AVZ 48.00 AVZ AR 120 2015 3 340 |
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10.1016/j.jbiosc.2015.01.007 doi PQ20160617 (DE-627)OLC1965022901 (DE-599)GBVOLC1965022901 (PRQ)g1424-5e69a1a7daf2db8085c07129bcb9aa3f95076b26af9c58682edf158fdc510f0f0 (KEY)0039398320150000120000300340efficientenrichmentofhighproducingrecombinantchine DE-627 ger DE-627 rakwb eng 570 540 660 DNB 42.00 bkl 58.30 bkl 48.00 bkl Okumura, Takeshi verfasserin aut Efficient enrichment of high-producing recombinant Chinese hamster ovary cells for monoclonal antibody by flow cytometry 2015 Text txt rdacontent ohne Hilfsmittel zu benutzen n rdamedia Band nc rdacarrier To screen a high-producing recombinant Chinese hamster ovary (CHO) cell from transfected cells is generally laborious and time-consuming. We developed an efficient enrichment strategy for high-producing cell screening using flow cytometry (FCM). A stable pool that had possibly shown a huge variety of monoclonal antibody (mAb) expression levels was prepared by transfection of an expression vector for mAb production to a CHO cell. To enrich high-producing cells derived from a stable pool stained with a fluorescent-labeled antibody that binds to mAb presented on the cell surface, we set the cell size and intracellular density gates based on forward scatter (FSC) and side scatter (SSC), and collected the brightest 5% of fluorescein isothiocyanate (FITC)-positive cells from each group by FCM. The final product concentration in a fed-batch culture of cells sorted without FSC and SSC gates was 1.2-1.3-times higher than that of unsorted cells, whereas that of cells gated by FSC and SSC was 3.4-4.7-fold higher than unsorted cells. Surprisingly, the fraction with the highest final product concentration indicated the smallest value of FSC and SSC, and the middle value of fluorescence intensity among all fractionated cells. Our results showed that our new screening strategy by FCM based on FSC and SSC gates could achieve an efficient enrichment of high-producing cells with the smallest value of FSC and SSC. Nutzungsrecht: Copyright © 2015 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved. © COPYRIGHT 2015 Elsevier B.V. Fluorescein Monoclonal antibodies Ovarian cancer Masuda, Kenji oth Watanabe, Kazuhiko oth Miyadai, Kenji oth Nonaka, Koichi oth Yabuta, Masayuki oth Omasa, Takeshi oth Enthalten in Journal of bioscience and bioengineering Osaka, 1999 120(2015), 3, Seite 340 (DE-627)265549817 (DE-600)1465387-4 (DE-576)074842005 1389-1723 nnns volume:120 year:2015 number:3 pages:340 http://dx.doi.org/10.1016/j.jbiosc.2015.01.007 Volltext http://www.ncbi.nlm.nih.gov/pubmed/25683450 GBV_USEFLAG_A SYSFLAG_A GBV_OLC SSG-OLC-TEC SSG-OLC-CHE SSG-OLC-FOR GBV_ILN_70 42.00 AVZ 58.30 AVZ 48.00 AVZ AR 120 2015 3 340 |
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10.1016/j.jbiosc.2015.01.007 doi PQ20160617 (DE-627)OLC1965022901 (DE-599)GBVOLC1965022901 (PRQ)g1424-5e69a1a7daf2db8085c07129bcb9aa3f95076b26af9c58682edf158fdc510f0f0 (KEY)0039398320150000120000300340efficientenrichmentofhighproducingrecombinantchine DE-627 ger DE-627 rakwb eng 570 540 660 DNB 42.00 bkl 58.30 bkl 48.00 bkl Okumura, Takeshi verfasserin aut Efficient enrichment of high-producing recombinant Chinese hamster ovary cells for monoclonal antibody by flow cytometry 2015 Text txt rdacontent ohne Hilfsmittel zu benutzen n rdamedia Band nc rdacarrier To screen a high-producing recombinant Chinese hamster ovary (CHO) cell from transfected cells is generally laborious and time-consuming. We developed an efficient enrichment strategy for high-producing cell screening using flow cytometry (FCM). A stable pool that had possibly shown a huge variety of monoclonal antibody (mAb) expression levels was prepared by transfection of an expression vector for mAb production to a CHO cell. To enrich high-producing cells derived from a stable pool stained with a fluorescent-labeled antibody that binds to mAb presented on the cell surface, we set the cell size and intracellular density gates based on forward scatter (FSC) and side scatter (SSC), and collected the brightest 5% of fluorescein isothiocyanate (FITC)-positive cells from each group by FCM. The final product concentration in a fed-batch culture of cells sorted without FSC and SSC gates was 1.2-1.3-times higher than that of unsorted cells, whereas that of cells gated by FSC and SSC was 3.4-4.7-fold higher than unsorted cells. Surprisingly, the fraction with the highest final product concentration indicated the smallest value of FSC and SSC, and the middle value of fluorescence intensity among all fractionated cells. Our results showed that our new screening strategy by FCM based on FSC and SSC gates could achieve an efficient enrichment of high-producing cells with the smallest value of FSC and SSC. Nutzungsrecht: Copyright © 2015 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved. © COPYRIGHT 2015 Elsevier B.V. Fluorescein Monoclonal antibodies Ovarian cancer Masuda, Kenji oth Watanabe, Kazuhiko oth Miyadai, Kenji oth Nonaka, Koichi oth Yabuta, Masayuki oth Omasa, Takeshi oth Enthalten in Journal of bioscience and bioengineering Osaka, 1999 120(2015), 3, Seite 340 (DE-627)265549817 (DE-600)1465387-4 (DE-576)074842005 1389-1723 nnns volume:120 year:2015 number:3 pages:340 http://dx.doi.org/10.1016/j.jbiosc.2015.01.007 Volltext http://www.ncbi.nlm.nih.gov/pubmed/25683450 GBV_USEFLAG_A SYSFLAG_A GBV_OLC SSG-OLC-TEC SSG-OLC-CHE SSG-OLC-FOR GBV_ILN_70 42.00 AVZ 58.30 AVZ 48.00 AVZ AR 120 2015 3 340 |
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10.1016/j.jbiosc.2015.01.007 doi PQ20160617 (DE-627)OLC1965022901 (DE-599)GBVOLC1965022901 (PRQ)g1424-5e69a1a7daf2db8085c07129bcb9aa3f95076b26af9c58682edf158fdc510f0f0 (KEY)0039398320150000120000300340efficientenrichmentofhighproducingrecombinantchine DE-627 ger DE-627 rakwb eng 570 540 660 DNB 42.00 bkl 58.30 bkl 48.00 bkl Okumura, Takeshi verfasserin aut Efficient enrichment of high-producing recombinant Chinese hamster ovary cells for monoclonal antibody by flow cytometry 2015 Text txt rdacontent ohne Hilfsmittel zu benutzen n rdamedia Band nc rdacarrier To screen a high-producing recombinant Chinese hamster ovary (CHO) cell from transfected cells is generally laborious and time-consuming. We developed an efficient enrichment strategy for high-producing cell screening using flow cytometry (FCM). A stable pool that had possibly shown a huge variety of monoclonal antibody (mAb) expression levels was prepared by transfection of an expression vector for mAb production to a CHO cell. To enrich high-producing cells derived from a stable pool stained with a fluorescent-labeled antibody that binds to mAb presented on the cell surface, we set the cell size and intracellular density gates based on forward scatter (FSC) and side scatter (SSC), and collected the brightest 5% of fluorescein isothiocyanate (FITC)-positive cells from each group by FCM. The final product concentration in a fed-batch culture of cells sorted without FSC and SSC gates was 1.2-1.3-times higher than that of unsorted cells, whereas that of cells gated by FSC and SSC was 3.4-4.7-fold higher than unsorted cells. Surprisingly, the fraction with the highest final product concentration indicated the smallest value of FSC and SSC, and the middle value of fluorescence intensity among all fractionated cells. Our results showed that our new screening strategy by FCM based on FSC and SSC gates could achieve an efficient enrichment of high-producing cells with the smallest value of FSC and SSC. Nutzungsrecht: Copyright © 2015 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved. © COPYRIGHT 2015 Elsevier B.V. Fluorescein Monoclonal antibodies Ovarian cancer Masuda, Kenji oth Watanabe, Kazuhiko oth Miyadai, Kenji oth Nonaka, Koichi oth Yabuta, Masayuki oth Omasa, Takeshi oth Enthalten in Journal of bioscience and bioengineering Osaka, 1999 120(2015), 3, Seite 340 (DE-627)265549817 (DE-600)1465387-4 (DE-576)074842005 1389-1723 nnns volume:120 year:2015 number:3 pages:340 http://dx.doi.org/10.1016/j.jbiosc.2015.01.007 Volltext http://www.ncbi.nlm.nih.gov/pubmed/25683450 GBV_USEFLAG_A SYSFLAG_A GBV_OLC SSG-OLC-TEC SSG-OLC-CHE SSG-OLC-FOR GBV_ILN_70 42.00 AVZ 58.30 AVZ 48.00 AVZ AR 120 2015 3 340 |
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Efficient enrichment of high-producing recombinant Chinese hamster ovary cells for monoclonal antibody by flow cytometry |
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To screen a high-producing recombinant Chinese hamster ovary (CHO) cell from transfected cells is generally laborious and time-consuming. We developed an efficient enrichment strategy for high-producing cell screening using flow cytometry (FCM). A stable pool that had possibly shown a huge variety of monoclonal antibody (mAb) expression levels was prepared by transfection of an expression vector for mAb production to a CHO cell. To enrich high-producing cells derived from a stable pool stained with a fluorescent-labeled antibody that binds to mAb presented on the cell surface, we set the cell size and intracellular density gates based on forward scatter (FSC) and side scatter (SSC), and collected the brightest 5% of fluorescein isothiocyanate (FITC)-positive cells from each group by FCM. The final product concentration in a fed-batch culture of cells sorted without FSC and SSC gates was 1.2-1.3-times higher than that of unsorted cells, whereas that of cells gated by FSC and SSC was 3.4-4.7-fold higher than unsorted cells. Surprisingly, the fraction with the highest final product concentration indicated the smallest value of FSC and SSC, and the middle value of fluorescence intensity among all fractionated cells. Our results showed that our new screening strategy by FCM based on FSC and SSC gates could achieve an efficient enrichment of high-producing cells with the smallest value of FSC and SSC. |
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To screen a high-producing recombinant Chinese hamster ovary (CHO) cell from transfected cells is generally laborious and time-consuming. We developed an efficient enrichment strategy for high-producing cell screening using flow cytometry (FCM). A stable pool that had possibly shown a huge variety of monoclonal antibody (mAb) expression levels was prepared by transfection of an expression vector for mAb production to a CHO cell. To enrich high-producing cells derived from a stable pool stained with a fluorescent-labeled antibody that binds to mAb presented on the cell surface, we set the cell size and intracellular density gates based on forward scatter (FSC) and side scatter (SSC), and collected the brightest 5% of fluorescein isothiocyanate (FITC)-positive cells from each group by FCM. The final product concentration in a fed-batch culture of cells sorted without FSC and SSC gates was 1.2-1.3-times higher than that of unsorted cells, whereas that of cells gated by FSC and SSC was 3.4-4.7-fold higher than unsorted cells. Surprisingly, the fraction with the highest final product concentration indicated the smallest value of FSC and SSC, and the middle value of fluorescence intensity among all fractionated cells. Our results showed that our new screening strategy by FCM based on FSC and SSC gates could achieve an efficient enrichment of high-producing cells with the smallest value of FSC and SSC. |
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To screen a high-producing recombinant Chinese hamster ovary (CHO) cell from transfected cells is generally laborious and time-consuming. We developed an efficient enrichment strategy for high-producing cell screening using flow cytometry (FCM). A stable pool that had possibly shown a huge variety of monoclonal antibody (mAb) expression levels was prepared by transfection of an expression vector for mAb production to a CHO cell. To enrich high-producing cells derived from a stable pool stained with a fluorescent-labeled antibody that binds to mAb presented on the cell surface, we set the cell size and intracellular density gates based on forward scatter (FSC) and side scatter (SSC), and collected the brightest 5% of fluorescein isothiocyanate (FITC)-positive cells from each group by FCM. The final product concentration in a fed-batch culture of cells sorted without FSC and SSC gates was 1.2-1.3-times higher than that of unsorted cells, whereas that of cells gated by FSC and SSC was 3.4-4.7-fold higher than unsorted cells. Surprisingly, the fraction with the highest final product concentration indicated the smallest value of FSC and SSC, and the middle value of fluorescence intensity among all fractionated cells. Our results showed that our new screening strategy by FCM based on FSC and SSC gates could achieve an efficient enrichment of high-producing cells with the smallest value of FSC and SSC. |
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Efficient enrichment of high-producing recombinant Chinese hamster ovary cells for monoclonal antibody by flow cytometry |
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