title: Comparative characterization of mAb producing hapten-specific hybridoma cells by flow cytometric analysis and ELISA.
abstract: 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 perform a high-throughput screening for hybridoma cells, which have both an adequate IgG production rate and a high target affinity.
authors: BAM Federal Institute for Materials Research and Testing, Richard-Willstätter-Str. 11, D-12489 Berlin, Germany.
authors: Maren
authors: M
authors: Kuhne
authors: BAM Federal Institute for Materials Research and Testing, Richard-Willstätter-Str. 11, D-12489 Berlin, Germany; University of Potsdam, Institute for Biochemistry and Biology, Karl-Liebknecht-Str. 24-25, 14476 Potsdam, Germany.
authors: Martin
authors: M
authors: Dippong
authors: BAM Federal Institute for Materials Research and Testing, Richard-Willstätter-Str. 11, D-12489 Berlin, Germany.
authors: Sabine
authors: S
authors: Flemig
authors: BAM Federal Institute for Materials Research and Testing, Richard-Willstätter-Str. 11, D-12489 Berlin, Germany.
authors: Katrin
authors: K
authors: Hoffmann
authors: BAM Federal Institute for Materials Research and Testing, Richard-Willstätter-Str. 11, D-12489 Berlin, Germany.
authors: Kristin
authors: K
authors: Petsch
authors: UP Transfer GmbH, Am Neuen Palais 10, D-14469 Potsdam, Germany; Hybrotec GmbH, Am Mühlenberg 11, D-14476 Potsdam-Golm, Germany.
authors: Jörg A
authors: JA
authors: Schenk
authors: BAM Federal Institute for Materials Research and Testing, Richard-Willstätter-Str. 11, D-12489 Berlin, Germany.
authors: Hans-Jörg
authors: HJ
authors: Kunte
authors: BAM Federal Institute for Materials Research and Testing, Richard-Willstätter-Str. 11, D-12489 Berlin, Germany. Electronic address: rudolf.schneider@bam.de.
authors: Rudolf J
authors: RJ
authors: Schneider
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000032:analysis* / Q000096:biosynthesis; D000915:Antibody Affinity; D000921:Antibody-Producing Cells / Q000276:immunology*; D000941:Antigens / Q000737:chemistry / Q000276:immunology; D002999:Clone Cells; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D004970:Estrone / Q000737:chemistry / Q000276:immunology; D005260:Female; D005434:Flow Cytometry / Q000379:methods; D006241:Haptens / Q000737:chemistry* / Q000276:immunology; D057166:High-Throughput Screening Assays; D006825:Hybridomas / Q000276:immunology*; D007074:Immunoglobulin G / Q000737:chemistry; D007150:Immunohistochemistry; D051379:Mice; D008807:Mice, Inbred BALB C; D012710:Serum Albumin, Bovine / Q000737:chemistry / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D000941:Antigens; D006241:Haptens; D007074:Immunoglobulin G; D012710:Serum Albumin, Bovine; D004970:Estrone
keywords: ELISA; Flow cytometry; Hapten; Hybridoma; Immunization; Monoclonal antibodies