CoChat
Sign Up
Cited by 34Monoclonal and Polyclonal Antibodies ResearchAnimal Genetics and ReproductionProtein purification and stability

A novel selection strategy for antibody producing hybridoma cells based on a new transgenic fusion cell line

Open access
Listek, Martin, Hönow, Anja, Gossen, Manfred, Hanack, Katja·2020·Scientific Reports
DOI 10.1038/s41598-020-58571-wPMC PMC6997400
Open this paper in CoChat — free accountOpen PDF

Abstract

The use of monoclonal antibodies is ubiquitous in science and biomedicine but the generation and validation process of antibodies is nevertheless complicated and time-consuming. To address these issues we developed a novel selective technology based on an artificial cell surface construct by which secreted antibodies were connected to the corresponding hybridoma cell when they possess the desired antigen-specificity. Further the system enables the selection of desired isotypes and the screening for potential cross-reactivities in the same context. For the design of the construct we combined the transmembrane domain of the EGF-receptor with a hemagglutinin epitope and a biotin acceptor peptide and performed a transposon-mediated transfection of myeloma cell lines. The stably transfected myeloma cell line was used for the generation of hybridoma cells and an antigen- and isotype-specific screening method was established. The system has been validated for globular protein antigens as well as for haptens and enables a fast and early stage selection and validation of monoclonal antibodies in one step.

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Integrity signals

Registry

Crossref · 10.1038/s41598-020-58571-w

Cites

2 of 2 references verified0 citation errors · 0 not checkable

Statistics

Claims the paper makes

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • Hybridomas are intentionally created fusions, typically between mouse myeloma cells and B-cells from various species, including humans.

    “Hybridomas present a unique challenge as they are intentionally created fusions, typically between mouse myeloma cells and B-cells from various species, including humans [71].”

    From Genetic Insights into the Economic Toll of Cell Line Misidentification: A Comprehensive Review (2026) · cites this paper as [71]

    The Introduction describes hybridomas formed by fusing myeloma cells with B lymphocytes, and the Methods identify the myeloma cells as murine. The study uses lymphocytes from mice; our copy does not support the added claim that the B cells typically come from various species, including humans. Page numbers were not supplied. (LLM, conf=0.88)

    Experimental — accuracy not yet measured

  • Murine hybridoma libraries provide monoclonal antibodies.

    “Murine hybridoma libraries provide monoclonal antibodies, insect cells power baculovirus expression systems, and plant protoplasts allow for CRISPR screens in agriculture [71,72,73].”

    From Genetic Insights into the Economic Toll of Cell Line Misidentification: A Comprehensive Review (2026) · cites this paper as [71]

  • Insect cells power baculovirus expression systems.

    “Murine hybridoma libraries provide monoclonal antibodies, insect cells power baculovirus expression systems, and plant protoplasts allow for CRISPR screens in agriculture [71,72,73].”

    From Genetic Insights into the Economic Toll of Cell Line Misidentification: A Comprehensive Review (2026) · cites this paper as [71]

  • Plant protoplasts allow for CRISPR screens in agriculture.

    “Murine hybridoma libraries provide monoclonal antibodies, insect cells power baculovirus expression systems, and plant protoplasts allow for CRISPR screens in agriculture [71,72,73].”

    From Genetic Insights into the Economic Toll of Cell Line Misidentification: A Comprehensive Review (2026) · cites this paper as [71]

    The Introduction and Conclusions discuss murine hybridomas and screening for antibody-producing cells, not plant protoplasts. A search of our copy for plant protoplasts, CRISPR screens, and agricultural genome editing found no passage addressing the attributed claim. (LLM, conf=0.77)

    Experimental — accuracy not yet measured

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

The self-check needs the paper’s full text.

Cites

Flagged first, then by citations

Research works printed but not linked: 38

  1. [1]Burton DR Wilson IA Immunology Square-dancing antibodies. Sci. 2007 317 5844 1507 8 17872431
  2. [2]Alkan SS Monoclonal antibodies: the story of a discovery that revolutionized science and medicine Nat. Rev. Immunol. 2004 4 2 153 6 10.1038/nri1265 15040588 · DOI
  3. [3]Geyer CR McCafferty J Dübel S Bradbury AR Sidhu SS Recombinant antibodies and in vitro selection technologies Methods Mol. Biol. 2012 901 11 32 10.1007/978-1-61779-931-0_2 22723092 · DOI
  4. [6]Reiss PD Min D Leung MY Working towards a consensus for antibody validation F1000Res 2014 3 266 2014 6 PMC4288440 25580232
  5. [7]Freedman LP The need for improved education and training in research antibody usage and validation practices Biotechniques 2016 61 1 16 8 10.2144/000114431 27401669 · DOI
  6. [8]Pauly D Hanack K How to avoid pitfalls in antibody use F1000Res 2015 4 691 10.12688/f1000research.6894.1 26834988 PMC4722690 · DOI
  7. [9]Zhang C Hybridoma technology for the generation of monoclonal antibodies Methods Mol. Biol. 2012 901 117 135 10.1007/978-1-61779-931-0_7 22723097 · DOI
  8. [10]Hanack K Messerschmidt K Listek M Antibodies and selection of monoclonal antibodies Adv. Exp. Med. Biol. 2016 917 11 22 10.1007/978-3-319-32805-8_2 27236550 · DOI
  9. [11]Köhler G Milstein C Continuous cultures of fused cells secreting antibody of predefined specificity Nat. 1975 256 5517 495 7 10.1038/256495a0 1172191 · DOI
  10. [12]Manz R Assenmacher M Pflüger E Miltenyi S Radbruch A Analysis and sorting of live cells according to secreted molecules, relocated to a cell-surface affinity matrix Proc. Natl Acad. Sci. USA 1995 92 6 1921 5 10.1073/pnas.92.6.1921 7892200 PMC42394 · DOI
  11. [13]Chen I Howarth M Lin W Ting AY Site-specific labeling of cell surface proteins with biophysical probes using biotin ligase Nat. Methods 2005 2 2 99 104 10.1038/nmeth735 15782206 · DOI
  12. [14]Zhen Y Caprioli RM Staros JV Characterization of glycosylation sites of the epidermal growth factor receptor Biochem. 2003 42 18 5478 92 10.1021/bi027101p 12731890 PMC2765783 · DOI
  13. [15]Jorisson RN Epidermal growth factor receptor: mechanisms of activation and signalling Exp. Cell Res. 2003 284 1 31 53 10.1016/S0014-4827(02)00098-8 12648464 · DOI · Paper page
  14. [16]He C Hobert M Friend L Carlin C The epidermal growth factor receptor juxtamembrane domain has multiple basolateral plasma membrane localization determinants, including a dominant signal with a polyproline core J. Biol. Chem. 2002 277 41 38284 93 10.1074/jbc.M104646200 12161422 · DOI
  15. [17]Lanzavecchia A Receptor-mediated uptake and its effect on antigen presentation to class II-restricted T lymphocytes Ann. Rev. Immunol. 1990 8 73 93 10.1146/annurev.iy.08.040190.004013 2188679 · DOI
  16. [18]Tomita M Tsumoto K Hybridoma technologies for antibody production Immunotherapy 2011 3 3 371 80 10.2217/imt.11.4 21395379 · DOI
  17. [19]Browne SM Al-Rubeai M Selection methods for high-producing mammalian cell lines Trends Biotechnol. 2007 25 9 425 432 10.1016/j.tibtech.2007.07.002 17659798 · DOI
  18. [20]Caroll S Al-Rubeai M The selection of high-producing cell lines using flow cytometry and cell sorting Expert. Opin. Biol. Ther. 2004 4 11 1821 9 10.1517/14712598.4.11.1821 15500410 · DOI
  19. [21]Dharshanan S Hung CS Screening and subcloning of high producer transfectomas using semisolid media and automated colony picker Methods Mol. Biol. 2014 1131 105 12 10.1007/978-1-62703-992-5_7 24515462 · DOI
  20. [22]Weaver JC McGrath P Adams S Gel microdrop technology for rapid isolation of rare and high producer cells Nat. Med. 1997 3 5 583 5 10.1038/nm0597-583 9142132 · DOI
  21. [23]El Debs B Utharala R Balyasnikova IV Griffiths AD Merten CA Functional single-cell hybridoma screening using droplet-based microfluidics Proc. Natl Acad. Sci. USA 2012 109 29 11570 5 10.1073/pnas.1204514109 22753519 PMC3406880 · DOI
  22. [24]Eyer K Single-cell deep phenotyping of IgG-secreting cells for high-resolution immune monitoring Nat. Biotechnol. 2017 35 10 977 982 10.1038/nbt.3964 28892076 · DOI
  23. [25]Parks DR Bryan VM Oi VT Herzenberg LA Antigen-specific identification and cloning of hybridomas with a fluorescence-activated cell sorter Proc. Natl Acad. Sci. USA 1979 76 4 1962 6 10.1073/pnas.76.4.1962 287037 PMC383513 · DOI
  24. [26]Mattanovich D Borth N Applications of cell sorting in biotechnology Microb. Cell Fact. 2006 5 12 10.1186/1475-2859-5-12 16551353 PMC1435767 · DOI
  25. [27]McKinney KL Dilwith R Belfort G Manipulation of heterogeneous hybridoma cultures for overproduction of monoclonal antibodies Biotechnol. Prog. 1991 7 5 445 54 10.1021/bp00011a010 1367993 · DOI
  26. [28]Kromenaker SJ Srienc F Cell cycle kinetics of the accumulation of heavy and light chain immunoglobulin proteins in a mouse hybridoma cell line Cytotechnology 1994 14 3 205 18 10.1007/BF00749617 7765591 · DOI
  27. [29]Charlet M Kromenaker SJ Srienc F Surface IgG content of murine hybridomas: Direct evidence for variation of antibody secretion rates during the cell cycle Biotechnol. Bioeng. 1995 47 5 535 40 10.1002/bit.260470505 18623432 · DOI
  28. [30]Meilhoc E Wittrup KD Bailey JE Application of flow cytometric measurement of surface IgG in kinetic analysis of monoclonal antibody synthesis and secretion by murine hybridoma cells J. Immunol. Methods 1989 121 2 167 74 10.1016/0022-1759(89)90157-9 2760464 · DOI
  29. [31]Pierce PW Engineered cell surface expression of membrane immunoglobulin as a means to identify monoclonal antibody-secreting hybridomas J. Immunol. Methods 2002 31 343 28 41 PMC2953942 19187782
  30. [32]Li C Siemasko K Clark MR Song W Cooperative interaction of Iga and Igb of the BCR regulates the kinetics and specificity of antigen targeting Int. Immunol. 2002 14 10 1179 1191 10.1093/intimm/14.10.1179 12356683 · DOI
  31. [33]Werner J Gossen M Modes of TAL effector-mediated repression Nucleic Acid. Res. 2014 42 21 13061 73 10.1093/nar/gku1124 25389273 PMC4245958 · DOI
  32. [34]Petersen TN Brunak S von Heijne G Nielsen H SignalP 4.0: discriminating signal peptides from transmembrane regions Nat. Methods 2011 8 10 785 6 10.1038/nmeth.1701 21959131 · DOI · Paper page
  33. [35]Tusnády GE Simon I Principles governing amino acid composition of integral membrane proteins: application to topology prediction J. Mol. Biol. 1998 283 2 489 506 10.1006/jmbi.1998.2107 9769220 · DOI
  34. [36]Tusnády GE Simon I The HMMTOP transmembrane topology prediction server Bioinforma. 2001 17 9 849 850 10.1093/bioinformatics/17.9.849 11590105 · DOI
  35. [37]Kuroda, H., Marino, M. P., Kutner, R. H. & Reiser, J. Production of lentiviral vectors in protein-free media. Curr Protoc Cell Biol, Chapter 26:Unit 26.8 (2011). 21400699
  36. [38]Cadinanos J Bradley A Generation of an inducible and optimized piggyBac transposon system Nucleic Acids Res. 2007 35 12 e87 10.1093/nar/gkm446 17576687 PMC1919496 · DOI
  37. [39]Micheel B The production and radioimmunoassay application of monoclonal antibodies to fluorescein isothiocyanate (FITC) J. Immunol. Methods 1988 111 1 89 94 10.1016/0022-1759(88)90063-4 3134489 · DOI
  38. [40]Holzlöhner, P. & Hanack, K. Generation of murine monoclonal antibodies by hybridoma technology. J Vis Exp119 (2017). PMC5407676 28117810

Cited by

Sorted by citations

Authors
Listek, M., Hönow, A., Gossen, M., & Hanack, K.
Affiliations
  • University of Potsdam (DE)
  • Berlin-Brandenburger Centrum für Regenerative Therapien (DE)
Corresponding author
Hanack, Katja
ORCID
Venue
Scientific Reports
Publisher
Springer Science and Business Media LLC
Published
2020-02-03
Volume
10
Issue
1
Pages
1664-1664
ISSN
2045-2322 (ISSN-L)
DOI
10.1038/s41598-020-58571-w
PMC
PMC6997400
OpenAlex
W3003339328
Cited by
34
References
2
Citations by year
2026: 7 · 2025: 6 · 2024: 4 · 2023: 4 · 2022: 4 · 2021: 8 · 2020: 1
FWCI
2.07
Type
journal article
Language
en
Open access
Licence
Locations
  • Scientific Reports (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion)
  • publish.UP (University of Potsdam) (submittedVersion, cc-by)
  • Universität Potsdam (cc-by)
Funders
Bundesministerium für Bildung und Forschung (03IP703X)
Topics
  • Health Sciences › Medicine › Radiology, Nuclear Medicine and Imaging › Monoclonal and Polyclonal Antibodies Research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Animal Genetics and Reproduction
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein purification and stability
Keywords
monoclonal antibody selection, hybridoma, hemagglutinin epitope, biotin acceptor peptide, myeloma cell lines, antibody validation
MeSH terms
Animals, Antibodies, Monoclonal, Antibody Specificity, Antibody-Producing Cells, Epitopes, Antigens, Cell Fusion, Hemagglutinins, Humans, Hybridomas, Multiple Myeloma, Ovalbumin, Recombinant Fusion Proteins, Transfection, Biotinylation, Cell Line, Tumor, Mice, ErbB Receptors

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

Are you an author, or do you see a problem with this page? Request a review