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Updated: Jan 26, 2026
Generation of Monoclonal Antibodies Against Natural Products
Published on: April 6, 2019
Immunochemical characterization of wild-type and variant glucocorticoid receptors by monoclonal antibodies
Abstract:
Monoclonal antibodies raised against the rat liver glucocorticoid receptor were used to investigate receptors of wild-type and glucocorticoid-resistant variants of mouse lymphoma cells. Two of the variant types contained receptors of 'nuclear transfer deficient' (nt-) and 'increased nuclear transfer' (nti) phenotypes, respectively, while the third was of the 'receptorless' (r-) phenotype with negligible hormone binding activity. Three monoclonal antibodies of the IgM class and one of the IgG class reacted with both wild-type and nt- receptors but not with the steroid binding form of nti receptors. Some of the antibodies bound the wild-type and nt- receptors more efficiently after activation at 20 degrees C. By use of an immuno-competition assay we were able to detect cross-reacting material in considerable amounts in extracts of nti and r- cell variants. This material was further characterized by gel filtration and immunoblotting. The immunoreactive material of wild-type, nti and r- cells gave a major band of mol. wt. 94 000 upon SDS-gel electrophoresis while the steroid-binding polypeptides of wild-type and nti receptors have mol. wts. of 94 000 and 40 000, respectively. The data show that in S49.1 mouse lymphoma cells the products of two receptor alleles can be distinguished.
Insights
Monoclonal antibodies reveal distinct forms of the glucocorticoid receptor in mouse lymphoma cells. This research differentiates receptor variants, aiding in understanding gene expression and drug resistance.
Area of Science:
- Molecular Biology
- Cell Biology
- Immunology
Background:
- Glucocorticoid receptors (GRs) regulate gene expression.
- Mutant mouse lymphoma cell lines exhibit varying GR functions, including altered nuclear transfer and hormone binding.
- Understanding GR variants is crucial for studying steroid hormone action and resistance.
Purpose of the Study:
- To characterize glucocorticoid receptors in wild-type and resistant mouse lymphoma cell variants using monoclonal antibodies.
- To investigate the structural and functional differences between GRs in different cell phenotypes.
- To identify cross-reacting material in receptor variants and distinguish receptor products.
Main Methods:
- Production and application of monoclonal antibodies against rat liver GR.
- Immuno-competition assays to detect cross-reacting GR material.
- SDS-gel electrophoresis, gel filtration, and immunoblotting for protein characterization.
Main Results:
- Monoclonal antibodies reacted with wild-type and 'nuclear transfer deficient' (nt-) receptors but not with the steroid-binding form of 'increased nuclear transfer' (nti) receptors.
- Cross-reacting material was detected in 'receptorless' (r-) and nti variants.
- Immunoreactive material in wild-type, nti, and r- cells showed a 94,000 mol. wt. band, distinct from the 40,000 mol. wt. steroid-binding polypeptide in nti receptors.
Conclusions:
- Monoclonal antibodies are effective tools for distinguishing GR variants.
- Mouse lymphoma cell variants possess distinct GR forms, including cross-reacting material.
- The study successfully differentiated products of two GR alleles in S49.1 mouse lymphoma cells.
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