The selection of high-producing cell lines using flow cytometry and cell sorting

Carroll, S M; Al‐Rubeai, Mohamed

doi:10.1517/14712598.4.11.1821

Cited by 76 publications

(39 citation statements)

References 51 publications

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“…It has been referred to as one of the main bottlenecks of getting a producer cell line out of development and into the manufacturing environment (Borth et al 2000;Carroll and Al-Rubeai 2004). A heterogeneous cell population that undergoes selection for high producers have the disadvantage that the high producing clones mainly have a reduced proliferation rate due to the energy being diverted towards production (Borth et al 2000;Cytotechnology (2007) 53:3-22 5 Kim and Lee 1999;Kromenaker and Srienc 1994).…”

Section: Optimization Of Selection Proceduresmentioning

confidence: 99%

“…This complex forms the primary means of capturing the secreted target antibody product which can then be fluorescently labeled for sorting by flow cytometry. This method significantly speeds up the selection process (Carroll and Al-Rubeai 2004), where the critical path activity of commercial cell lines can be minimized to 31 weeks compared to a conventional 40 (Racher 2006). …”

Section: Optimization Of Selection Proceduresmentioning

confidence: 99%

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Using cell engineering and omic tools for the improvement of cell culture processes

et al. 2007

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Significant strides have been made in mammalian cell based biopharmaceutical process and cell line development over the past years. With several established mammalian host cell lines and expression systems, optimization of selection systems to reduce development times and improvement of glycosylation patterns are only some of the advances being made to improve cell culture processes. In this article, the advances pertaining to cell line development and cell engineering strategies are discussed. An overview of the cell engineering strategies to enhance cellular characteristics by genetic manipulation are illustrated, focusing on the use of genomics and proteomics tools and their application in such endeavors. Included in this review are some of the early studies using the 'omic' technique to understand cellular mechanisms of product synthesis and secretion, apoptosis, cell proliferation and the influence of the physicochemical environment. The article highlights the significance of integrating genomics and proteomics data with the vast amounts of bioprocess data for improved analysis of the biological pathways involved. Further improvements of the techniques and methodologies used are needed but ultimately, the new cell engineering strategies should provide great insight into the regulatory networks within the cell in a bioprocess environment and how to manipulate them to increase overall productivity.

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Section: Optimization Of Selection Proceduresmentioning

confidence: 99%

Section: Optimization Of Selection Proceduresmentioning

confidence: 99%

Using cell engineering and omic tools for the improvement of cell culture processes

et al. 2007

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“…The successful development of producer cell lines is a long and strenuous procedure with hundreds to thousands of potential clones screened, tested and thoroughly characterized. Methods for cell line development rely on classical, dilution‐based plating as well as high throughput methods using robotics or fluorescence activated cell sorting (FACS) 2, 3 followed by detailed characterization of subclones. Moreover, testing the identity and stability of producer lines from master to working cell bank to process is complex and time consuming.…”

Section: Introductionmentioning

confidence: 99%

Label‐free live cell imaging by Confocal Raman Microscopy identifies CHO host and producer cell lines

Mateu

Harreither

Schosserer

et al. 2016

Biotechnology Journal

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As a possible viable and non‐invasive method to identify high producing cells, Confocal Raman Microscopy was shown to be able to differentiate CHO host cell lines and derivative production clones. Cluster analysis of spectra and their derivatives was able to differentiate between different producer cell lines and a host, and also distinguished between an intracellular region of high lipid and protein content that in structure resembles the Endoplasmic Reticulum. This ability to identify the ER may be a major contributor to the identification of high producers. PCA enabled the discrimination even of host cell lines and their subclones with inherently higher production capacity. The method is thus a promising option that may contribute to early, non‐invasive identification of high potential candidates during cell line development and possibly could also be used for proof of identity of established production clones.

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“…A distribution curve of productivity of clones, originating from one transfection event, usually shows only a few clones that have significantly higher productivity levels. In most cases, the number of high producing clones detected depend on the number of clones screened (Carroll and Al-Rubeai 2004). The clone selection process has been referred to as the main bottleneck when generating a high producing cell line, since sub cloning and testing is usually time consuming and labour intensive.…”

Section: Introductionmentioning

confidence: 99%

“…Several techniques have been developed with the intention to reduce time constraints and manpower costs during cell line development. This includes fluorescence based systems such as fluorescence-activated cell sorting, FACS (Carroll and Al-Rubeai 2004;Brezinsky et al 2003;Sleiman et al 2007), and more recently, the Genetix ClonePix FL system. We have chosen another approach to reduce cost and decrease labour, which is not dependent on fluorescence technology.…”

Section: Introductionmentioning

confidence: 99%

Automation of cell line development

et al. 2009

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An automated platform for development of high producing cell lines for biopharmaceutical production has been established in order to increase throughput and reduce development costs. The concept is based on the Cello robotic system (The Automation Partnership) and covers screening for colonies and expansion of static cultures. In this study, the glutamine synthetase expression system (Lonza Biologics) for production of therapeutic monoclonal antibodies in Chinese hamster ovary cells was used for evaluation of the automation approach. It is shown that the automated procedure is capable of producing cell lines of equal quality to the traditionally generated cell lines in terms of colony detection following transfection and distribution of IgG titer in the screening steps. In a generic fed-batch evaluation in stirred tank bioreactors, IgG titers of 4.7 and 5.0 g/L were obtained for best expressing cell lines. We have estimated that the number of completed cell line development projects can be increased up to three times using the automated process without increasing manual workload, compared to the manual process. Correlation between IgG titers obtained in early screens and titers achieved in fed-batch cultures in shake flasks was found to be poor. This further implies the benefits of utilizing a high throughput system capable of screening and expanding a high number of transfectants. Two concentrations, 56 and 75 lM, of selection agent, methionine sulphoximine (MSX), were applied to evaluate the impact on the number of colonies obtained post transfection. When applying selection medium containing 75 lM MSX, fewer low producing transfectants were obtained, compared to cell lines selected with 56 lM MSX, but an equal number of high producing cell lines were found. By using the higher MSX concentration, the number of cell line development projects run in parallel could be increased and thereby increasing the overall capacity of the automated platform process.

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The selection of high-producing cell lines using flow cytometry and cell sorting

Cited by 76 publications

References 51 publications

Using cell engineering and omic tools for the improvement of cell culture processes

Using cell engineering and omic tools for the improvement of cell culture processes

Label‐free live cell imaging by Confocal Raman Microscopy identifies CHO host and producer cell lines

Automation of cell line development

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