Production and characterization of monoclonal antibody against a triple negative breast cancer cell line
- Department of Immunology, School of Medicine, Shiraz University of Medical Sciences, Shiraz (Iran, Islamic Republic of)
- Shiraz Institute for Cancer Research, School of Medicine, Shiraz University of Medical Sciences, Shiraz (Iran, Islamic Republic of)
- Fellow of Molecular Pathology and Cytogenetics, Department of Pathology, School of Medicine, Shiraz University of Medical Sciences, Shiraz (Iran, Islamic Republic of)
Highlights: • Discovery of potential biomarkers that can predict tumor behavior is essential in breast cancer. • A panel of monoclonal antibodies was produced by hybridoma technology. • MAbs that reacted with surface antigens of immunizing cell line but not normal cells were selected. • Target was identified as NCAM1 by immunoprecipitation and mass spectrometry. • The anti-NCAM1 antibody has potential application in cancer diagnosis. Breast cancer is the most prevalent malignancy among women around the world such that more than 1,400,000 new cases are being diagnosed each year. Despite immense studies over many years on diagnosis and treatment of breast cancer, about 30% of treated patients will relapse and require subsequent therapy. By development of hybridoma technology, murine monoclonal antibodies (MAbs) against several human tumor-associated antigens have been produced and characterized in many laboratories. The purpose of these studies is to generate effective monoclonal antibodies that could be useful in tumor diagnosis and therapy. In this study, splenic lymphocytes of immunized BALB/c mouse with a new established breast cancer cell line (Pari-ICR cell line, established in Shiraz Institute for Cancer Research) were fused with the mouse myeloma cell line SP2/0 in the presence of polyethylene glycol. We generated a panel of monoclonal antibodies against the newly established cell line. The hybrid cultures were screened by flow cytometry. Hybridomas that produced antibody to surface antigens of immunizing cell line but not to Human Gingival Fibroblasts, adipose stem cells, and leucocytes isolated from peripheral blood were selected and cloned by limiting dilution method. The 1E3 clone (IgG2a type) that displayed clonal stability was further analyzed for specificity by flow cytometry. MAb 1E3 showed weak to strong reactivity to other cell lines compared with Pari-ICR cell line. Antigen identification was performed by a workflow consisting of immunoaffinity purification, SDS-PAGE, Western blotting, and mass spectrometry analysis. The target of 1E3 mAb was identified as NCAM1. In conclusion, using the antibody-based strategy we identified NCAM1 as a potential therapeutic target and biomarker for breast cancer.
- OSTI ID:
- 23107830
- Journal Information:
- Biochemical and Biophysical Research Communications, Vol. 505, Issue 1; Other Information: Copyright (c) 2018 Elsevier Inc. All rights reserved.; Country of input: International Atomic Energy Agency (IAEA); ISSN 0006-291X
- Country of Publication:
- United States
- Language:
- English
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