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NCL-CB11, a new monoclonal antibody recognizing the internal domain of the c-erbB-2 oncogene protein effective for
I P Corbett1, J A Henry, B Angus
1Department of Pathology, University of Newcastle upon Tyne, U.K.
Abstract:
The c-erbB-2 proto-oncogene encodes a 190 k Mr protein representing a putative growth factor receptor with considerable homology to the EGF receptor. Gene amplification and overexpression of the oncogene protein have been demonstrated in a variety of tumours including breast cancer, where it is associated with a poor prognosis. In this study we have produced and characterized a mouse monoclonal antibody, designated NCL-CB11, to the c-erbB-2 protein. NCL-CB11 was generated to a synthetic peptide sequence corresponding to a site of predicted antigenicity near the C-terminus of the internal domain of the protein. NCL-CB11 produces intense membrane-associated immunohistochemical staining in a proportion of human cancer cells. The specificity of the antibody is supported by Western blotting and immunoprecipitation studies. Reactivity with an internal site of the protein is confirmed by the necessity of cell permeabilization for reactivity in fluorescence-activated cell sorter (FACS) analysis. A high degree of correlation between immunohistochemical staining using NCL-CB11, and c-erbB-2 gene amplification has been observed. NCL-CB11 should prove to be a valuable reagent for investigations into the pathological significance of c-erbB-2 protein expression.
Insights
Researchers developed a new antibody, NCL-CB11, targeting the c-erbB-2 oncoprotein. This antibody shows promise for detecting c-erbB-2 protein expression in tumors, aiding in cancer research and diagnosis.
Area of Science:
- Oncology
- Molecular Biology
- Immunology
Background:
- The c-erbB-2 proto-oncogene encodes a protein homologous to the EGF receptor.
- Amplification and overexpression of c-erbB-2 are linked to poor prognosis in various cancers, including breast cancer.
Purpose of the Study:
- To produce and characterize a novel mouse monoclonal antibody, NCL-CB11, against the c-erbB-2 protein.
- To evaluate the antibody's utility in detecting c-erbB-2 protein expression in human tumors.
Main Methods:
- Generation of a monoclonal antibody (NCL-CB11) using a synthetic peptide near the C-terminus of the c-erbB-2 protein.
- Immunohistochemical staining, Western blotting, immunoprecipitation, and fluorescence-activated cell sorter (FACS) analysis were used for characterization.
- Correlation analysis between NCL-CB11 staining and c-erbB-2 gene amplification.
Main Results:
- NCL-CB11 demonstrated intense, membrane-associated immunohistochemical staining in a subset of human cancer cells.
- Specificity was confirmed by Western blotting and immunoprecipitation.
- FACS analysis confirmed reactivity with an internal protein site requiring cell permeabilization.
- A strong correlation was observed between NCL-CB11 immunohistochemical staining and c-erbB-2 gene amplification.
Conclusions:
- The NCL-CB11 antibody is a specific and valuable reagent for detecting c-erbB-2 protein.
- This antibody can be used to investigate the pathological significance of c-erbB-2 protein expression in cancer.