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Expression of a recombinant sheep IgE gene
1CSIRO Division of Animal Health, McMaster Laboratory, NSW, Australia.
Immunological Investigations
|January 1, 1994
Summary
Researchers generated novel ovine-murine immunoglobulin E (IgE) monoclonal antibodies using advanced molecular genetics. This breakthrough facilitates the study of low-abundance immunoglobulins and enhances immunological research capabilities.
Area of Science:
- Immunology
- Molecular Genetics
- Biochemistry
Background:
- Immunoglobulin E (IgE) plays a crucial role in allergic reactions and parasite defense.
- Studying IgE is challenging due to its low abundance in serum.
- Development of specific reagents is vital for IgE research.
Purpose of the Study:
- To develop a method for producing monoclonal antibodies against ovine IgE.
- To utilize molecular genetic techniques for generating specific immunological reagents.
- To facilitate further immunological studies by enabling monoclonal antibody production.
Main Methods:
- Screening an ovine cosmid library with a bovine IgE constant region (C epsilon) gene probe.
- Isolating ovine C epsilon and IgA (C alpha) genes.
- Cloning the C epsilon gene into an expression vector with a specific variable heavy (VDJ) chain.
- Transfecting the construct into murine hybridoma cells.
- Affinity purification and characterization of chimeric ovine-murine IgE antibodies.
Main Results:
- Successfully isolated ovine IgE and IgA constant region genes.
- Generated stable transfectomas secreting chimeric ovine-murine IgE anti-NIP antibodies.
- Characterized the chimeric antibodies, confirming the presence of expected heavy and light chains.
Conclusions:
- Advanced molecular genetic techniques can successfully generate immunological reagents for low-abundance immunoglobulin isotypes like IgE.
- This methodology enables the production of specific monoclonal antibodies, advancing immunological research.
- The developed chimeric antibodies serve as valuable tools for studying IgE functions.
