Abstract
A murine hybridoma secreting monoclonal IgE antibodies of anti-2,4-dinitrophenyl (DNP) specificity was generated by fusion of SP2/0 tumor cells and spleen cells from DNP-Ascaris-hyperimmunized mice. Hybridomas secreting anti-DNP antibodies of other heavy chain classes, i.e., μ, γ 1 and γ(2b), were also obtained from the same fusion experiment. Large quantities of IgE antibodies were obtained from ascites of mice in which the IgE-secreting hybridoma was propagated in vivo. The IgE antibodies were isolated by precipitation with ammonium sulfate followed by affinity chromatography on DNP-bovine serum albumin (BSA)-Sepharose-4B and further purified by ion-exchange chromatography on DEAE-cellulose and gel filtration on Sephadex G-200. The isolated IgE has an approximate m.w. of 184,000, a total carbohydrate content of 13.3%, and its amino acid composition has been determined. The antibody has an association constant with DNP-lysine of 1.4 x 10 8 M -1 at 25°C and 10 7 7 M -1 at 37°C. Rabbit and goat antibodies against the hybridoma IgE were prepared and the antisera were made specific for IgE by adsorption on normal mouse serum protein-Sepharose-4B. Solid phase radioimmunoassays for measuring murine antigen-specific and total IgE were developed and were shown to have high specificity and sensitivity. Finally, the isolated hybridoma IgE can mediate antigen (DNP-BSA)-induced release of mediator (serotonin) from rat basophilic leukemia cells.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 2728-2737 |
| Number of pages | 10 |
| Journal | Journal of Immunology |
| Volume | 124 |
| Issue number | 6 |
| State | Published - 1980 |
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ASJC Scopus subject areas
- Immunology
Cite this
Monoclonal dinitrophenyl-specific murine IgE antibody : Preparation, isolation, and characterization. / Liu, Fu-Tong; Bohn, J. W.; Ferry, E. L.; Yamamoto, H.; Molinaro, C. A.; Sherman, L. A.; Klinman, N. R.; Katz, D. H.
In: Journal of Immunology, Vol. 124, No. 6, 1980, p. 2728-2737.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Monoclonal dinitrophenyl-specific murine IgE antibody
T2 - Preparation, isolation, and characterization
AU - Liu, Fu-Tong
AU - Bohn, J. W.
AU - Ferry, E. L.
AU - Yamamoto, H.
AU - Molinaro, C. A.
AU - Sherman, L. A.
AU - Klinman, N. R.
AU - Katz, D. H.
PY - 1980
Y1 - 1980
N2 - A murine hybridoma secreting monoclonal IgE antibodies of anti-2,4-dinitrophenyl (DNP) specificity was generated by fusion of SP2/0 tumor cells and spleen cells from DNP-Ascaris-hyperimmunized mice. Hybridomas secreting anti-DNP antibodies of other heavy chain classes, i.e., μ, γ 1 and γ(2b), were also obtained from the same fusion experiment. Large quantities of IgE antibodies were obtained from ascites of mice in which the IgE-secreting hybridoma was propagated in vivo. The IgE antibodies were isolated by precipitation with ammonium sulfate followed by affinity chromatography on DNP-bovine serum albumin (BSA)-Sepharose-4B and further purified by ion-exchange chromatography on DEAE-cellulose and gel filtration on Sephadex G-200. The isolated IgE has an approximate m.w. of 184,000, a total carbohydrate content of 13.3%, and its amino acid composition has been determined. The antibody has an association constant with DNP-lysine of 1.4 x 10 8 M -1 at 25°C and 10 7 7 M -1 at 37°C. Rabbit and goat antibodies against the hybridoma IgE were prepared and the antisera were made specific for IgE by adsorption on normal mouse serum protein-Sepharose-4B. Solid phase radioimmunoassays for measuring murine antigen-specific and total IgE were developed and were shown to have high specificity and sensitivity. Finally, the isolated hybridoma IgE can mediate antigen (DNP-BSA)-induced release of mediator (serotonin) from rat basophilic leukemia cells.
AB - A murine hybridoma secreting monoclonal IgE antibodies of anti-2,4-dinitrophenyl (DNP) specificity was generated by fusion of SP2/0 tumor cells and spleen cells from DNP-Ascaris-hyperimmunized mice. Hybridomas secreting anti-DNP antibodies of other heavy chain classes, i.e., μ, γ 1 and γ(2b), were also obtained from the same fusion experiment. Large quantities of IgE antibodies were obtained from ascites of mice in which the IgE-secreting hybridoma was propagated in vivo. The IgE antibodies were isolated by precipitation with ammonium sulfate followed by affinity chromatography on DNP-bovine serum albumin (BSA)-Sepharose-4B and further purified by ion-exchange chromatography on DEAE-cellulose and gel filtration on Sephadex G-200. The isolated IgE has an approximate m.w. of 184,000, a total carbohydrate content of 13.3%, and its amino acid composition has been determined. The antibody has an association constant with DNP-lysine of 1.4 x 10 8 M -1 at 25°C and 10 7 7 M -1 at 37°C. Rabbit and goat antibodies against the hybridoma IgE were prepared and the antisera were made specific for IgE by adsorption on normal mouse serum protein-Sepharose-4B. Solid phase radioimmunoassays for measuring murine antigen-specific and total IgE were developed and were shown to have high specificity and sensitivity. Finally, the isolated hybridoma IgE can mediate antigen (DNP-BSA)-induced release of mediator (serotonin) from rat basophilic leukemia cells.
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UR - http://www.scopus.com/inward/citedby.url?scp=0018823631&partnerID=8YFLogxK
M3 - Article
C2 - 7373045
AN - SCOPUS:0018823631
VL - 124
SP - 2728
EP - 2737
JO - Journal of Immunology
JF - Journal of Immunology
SN - 0022-1767
IS - 6
ER -