Comparative characterization of mAb producing hapten-specific hybridoma cells by flow cytometric analysis and ELISA

  • A novel method that optimizes the screening for antibody-secreting hapten-specific hybridoma cells by using flow cytometry is described. Cell clones specific for five different haptens were analyzed. We selectively double stained and analyzed fixed hybridoma cells with fluorophore-labeled haptens to demonstrate the target-selectivity, and with a fluorophore-labeled anti-mouse IgG antibody to characterize the level of surface expression of membrane-bound IgGs. ELISA measurements with the supernatants of the individual hybridoma clones revealed that antibodies from those cells, which showed the highest fluorescence intensities in the flow cytometric analysis, also displayed the highest affinities for the target antigens. The fluorescence intensity of antibody-producing cells corresponded well with the produced antibodies' affinities toward their respective antigens. Immunohistochemical staining verified the successful double labeling of the cells. Our method makes it possible to performA novel method that optimizes the screening for antibody-secreting hapten-specific hybridoma cells by using flow cytometry is described. Cell clones specific for five different haptens were analyzed. We selectively double stained and analyzed fixed hybridoma cells with fluorophore-labeled haptens to demonstrate the target-selectivity, and with a fluorophore-labeled anti-mouse IgG antibody to characterize the level of surface expression of membrane-bound IgGs. ELISA measurements with the supernatants of the individual hybridoma clones revealed that antibodies from those cells, which showed the highest fluorescence intensities in the flow cytometric analysis, also displayed the highest affinities for the target antigens. The fluorescence intensity of antibody-producing cells corresponded well with the produced antibodies' affinities toward their respective antigens. Immunohistochemical staining verified the successful double labeling of the cells. Our method makes it possible to perform a high-throughput screening for hybridoma cells, which have both an adequate IgG production rate and a high target affinity.zeige mehrzeige weniger

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Metadaten
Autoren/innen:Maren Kuhne, Martin Dippong, Sabine Flemig, Katrin Hoffmann, K. Petsch, J.A. Schenk, Hans-Jörg Kunte, Rudolf Schneider
Dokumenttyp:Zeitschriftenartikel
Veröffentlichungsform:Verlagsliteratur
Sprache:Englisch
Titel des übergeordneten Werkes (Englisch):Journal of immunological methods
Jahr der Erstveröffentlichung:2014
Organisationseinheit der BAM:1 Analytische Chemie; Referenzmaterialien
1 Analytische Chemie; Referenzmaterialien / 1.8 Umweltanalytik
1 Analytische Chemie; Referenzmaterialien / 1.2 Biophotonik
4 Material und Umwelt
4 Material und Umwelt / 4.1 Biologische Materialschädigung und Referenzorganismen
Verlag:Elsevier B.V.
Verlagsort:Amsterdam
Jahrgang/Band:413
Erste Seite:45
Letzte Seite:56
Freie Schlagwörter:Aflatoxin; CLSM; Digoxigenin; ELISA; Estradiol; Estrone; Flow cytometry; Hapten; Hybridoma; Immunization; Monoclonal antibodies; Zearalenone
DOI:https://doi.org/10.1016/j.jim.2014.07.004
ISSN:0022-1759
ISSN:1872-7905
Verfügbarkeit des Dokuments:Datei im Netzwerk der BAM verfügbar ("Closed Access")
Datum der Freischaltung:20.02.2016
Referierte Publikation:Ja
Datum der Eintragung als referierte Publikation:22.12.2014