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Monoclonal antibodies to murine retrovirus protein p30
Abstract:
Hybrid cell lines were prepared by the fusion of mouse myeloma cells with the spleen cells of Wistar-Furth rats that had been immunized with a Moloney sarcoma virus (Mo-MuSV)-induced tumour, MFU. Two immunization protocols were designed. In the first, the animals received several injections of irradiated (10 000 rad) cells of a tumour cell line established in vitro, MFU-67. The rats received a booster injection 3 days prior to fusion. In the second protocol, immunization was the result of simple tumour growth, and no booster was given. Hybrids were tested by immunofluorescence for the production of immunoglobulins reacting with mouse cells acutely infected with Mo-MuSV. Over 20% of reactive hybrids were observed in the tumour growth protocol, and about 10% in the irradiated cell protocol when the last injection of the series was given 2 weeks before fusion. After 6 months, the proportion fell to 3%. Hybrid lines producing antibody to p30, the major core polypeptide of murine retroviruses, were obtained by cloning. Three of these were selected for closer study and were found to recognize three non-overlapping epitopes on p30. By direct and competitive binding in ELISA tests, the three epitopes were found to have very different distribution patterns among the various strains and isolates of murine retroviruses.
Insights
Researchers developed hybrid cell lines to create antibodies against Moloney sarcoma virus (Mo-MuSV). They identified antibodies targeting specific epitopes on the p30 protein of murine retroviruses.
Area of Science:
- Immunology
- Virology
- Hybridoma Technology
Background:
- Moloney sarcoma virus (Mo-MuSV) induces tumors, necessitating the development of targeted immunotherapies.
- Hybridoma technology offers a method for producing monoclonal antibodies against viral components.
Purpose of the Study:
- To generate hybrid cell lines producing antibodies against Mo-MuSV-induced tumors.
- To characterize antibodies targeting the p30 protein of murine retroviruses.
Main Methods:
- Fusion of mouse myeloma cells with spleen cells from immunized Wistar-Furth rats.
- Immunization using irradiated tumor cells or tumor growth protocols.
- Screening of hybrid cells via immunofluorescence for reactivity with Mo-MuSV-infected cells.
- Cloning of hybrid lines producing antibodies to the p30 protein.
- Epitope mapping using ELISA to determine antibody binding sites on p30.
Main Results:
- Two immunization protocols yielded reactive hybridomas, with the tumor growth protocol showing higher initial reactivity (>20%).
- Antibody-producing hybrid lines targeting the p30 protein were successfully generated.
- Three distinct, non-overlapping epitopes on the p30 protein were identified.
- ELISA revealed diverse distribution patterns of these epitopes across different murine retrovirus strains.
Conclusions:
- Hybridoma technology is effective for generating antibodies against Mo-MuSV-related antigens.
- The identified antibodies recognize specific epitopes on the p30 protein, aiding in the characterization of murine retroviruses.
- Understanding epitope distribution is crucial for developing broadly reactive diagnostic and therapeutic agents.