Clonal Stability and Heterogeneity of Hybridomas: Analysis by Multiparameter Flow Cytometry

Andreeff, Michael; Bartal, Arie H.; Feit, Carl; Hirshaut, Yashar

doi:10.1089/hyb.1985.4.277

Cited by 14 publications

(8 citation statements)

References 6 publications

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“…Also, the early detection of a particular class or subclass can contribute to the assessment of population dynamics involved in prolonged culture. (17) A potential disadvantage of the multiplexed bead analysis of antibody isotype is that signal isolation depends upon the quality of the capture reagent. Because flow cytometry histograms can be presented on a linear scale, significant cross-reactivity can produce visually similar histograms.…”

Section: Discussionmentioning

confidence: 99%

Structural Mapping of Immunoglobulin Subclasses Using Multiplexed Bead Flow Cytometry

et al. 2006

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Monoclonal hybridomas secrete immunoglobulin molecules with a single specificity and distinct class/subclass structure. The determination of immunoglobulin structure can be used to facilitate hybridoma colony management and predict monoclonality. In this report, we used multiplexed bead flow cytometry to define hybridoma class/subclass. The assay was sufficiently sensitive to detect 50 ng/mL of antibody. The multiplexed bead assay efficiently defined traditional class/subclass determinants as well as more subtle patterns of crossreactivity. Further, the assay was combined with Poisson statistics to provide a numerical estimate of hybridoma monoclonality. The sensitivity and flexibility of this approach suggests the utility of multiplexed bead flow cytometry in the early management of immunoglobulin-secreting hybridomas.

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Section: Discussionmentioning

confidence: 99%

Structural Mapping of Immunoglobulin Subclasses Using Multiplexed Bead Flow Cytometry

et al. 2006

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“…Supporting evidence for the monoclonality of our hybridomas is the presence of a single isotype and monospecificity by immunofluorescence; nonetheless, low frequency or non-producing contaminants cannot be excluded. An additional advantage of the alginate matrix is the ease with which repeat cloning can be performed in the presence of hybridoma instability (1). We have used the alginate matrix to clone established hybridomas to a variety of cell surface antigens.…”

Section: Discussionmentioning

confidence: 99%

“…A major limitation in current monoclonal antibody (MAb) production is the loss of antibody-producing hybridomas due to overgrowth by irrelevant or nonproducing hybridoma cells (1)(2)(3). The problem of hybridoma population dynamics has been addressed using several approaches.…”

Section: Introductionmentioning

confidence: 99%

Cloning Hybridomas in a Reversible Three-Dimensional Alginate Matrix

Li¹,

Abdi²,

Mentzer³

1992

Hybridoma

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Alginate is a transparent polymer of guluronic and mannuronic acids that provide a favorable microenvironment for cell growth. Alginate gelation is calcium dependent and temperature independent. To facilitate the isolation of stable and productive antibody-producing hybridomas, we have developed a technique of cloning hybridomas in the three-dimensional alginate matrix. To provide cavities for hybridoma growth, we encorporated 10-15% (v/v) gelatin into the alginate prior to gelation. We have cloned more than 90 monoclonal antibody-producing hybridomas using the alginate matrix. The alginate matrix is readily reversible with the addition of a calcium chelator. The alginate matrix permits efficient cloning in limited incubator space, without the use of a feeder layer, and with minimal amount of medium. The transparent matrix also permits easy screening for clonality and growth.

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“…Since Zhang's 2012 review, new technologies have become available to researchers that stand to take the hybridoma platform out of the dark ages: HyLiTE software, gold nano-particle laser cell fusion and distortion-based microfl uidics. Zhang's review focused on 40-year-old PEG technology for the fusion steps and standard techniques for screening (ELISA, fl ow cytometry, fl uorescence-activated cell sorting, and immunohistochemistry) that have been in use since the early 1980s [21][22][23][24]. Deformity-based microfl uidic fusion methods of fusion [19] and gold nano-particle laser cell fusion techniques [18], available only since 2014, are vast improvements over PEG fusion, possibly raising effi ciency rates for viable hybridomas from ∼1 in 10,000 cells to ∼9,500 in 10,000 cells [14,19].…”

Section: Trends That May Revolutionize the Hybridoma Platformmentioning

confidence: 99%

Life begins at forty – hybridomas: ageing technology holds promise for future drug discoveries

Steele¹

2016

GaBI J

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Hybridomas are used to produce monoclonal antibodies used in the prevention, diagnosis and treatment of diseases such as cancer, multiple sclerosis, heart disease, infl ammatory diseases, macular degeneration, transplant rejection, and viral infections [3]. Other systems for producing monoclonal antibodies have been developed since Milstein's pioneering work. Monoclonal anti bodies (mAbs) are also made using Chinese Hamster Ovaries (CHO), yeasts, insects, plants, bacteria and phages [3]. CHO is the most popular method of mAb production due to cheaper costs, favourable approval history and reliability in the lab [3]. Hybridomas could soon see increased use in mAb production as improvements are made to the hybridoma platform [4-6], see Table 1. Since their initial creation, hybridomas have been widely employed in the commercial production of mAbs. Initially, the main diffi culty with hybridomas was in generating highly

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Clonal Stability and Heterogeneity of Hybridomas: Analysis by Multiparameter Flow Cytometry

Cited by 14 publications

References 6 publications

Structural Mapping of Immunoglobulin Subclasses Using Multiplexed Bead Flow Cytometry

Structural Mapping of Immunoglobulin Subclasses Using Multiplexed Bead Flow Cytometry

Cloning Hybridomas in a Reversible Three-Dimensional Alginate Matrix

Life begins at forty – hybridomas: ageing technology holds promise for future drug discoveries

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