Production and Characterization of a Peptide-based Monoclonal Antibody Against CD44 Variant 6
| Autor: | Ali Ahmad Bayat, Mahmood Jeddi-Tehrani, Banafsheh Tavangar, Yasaman Vojgani, Saeed Zarei, Zahra Amirghofran, Ahmad Reza Mahmoudi, Reza Hadavi |
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| Rok vydání: | 2015 |
| Předmět: |
Immunoconjugates
medicine.drug_class Blotting Western Immunology Immunocytochemistry Enzyme-Linked Immunosorbent Assay Monoclonal antibody Cell Line Mice Antigen Affinity chromatography Western blot Antibody Specificity Antigens Neoplasm Cell Line Tumor medicine Animals Humans Immunology and Allergy Mice Inbred BALB C Hybridomas biology medicine.diagnostic_test CD44 Antibodies Monoclonal Original Articles Flow Cytometry Immunohistochemistry Molecular biology Founder Effect Protein Structure Tertiary Hyaluronan Receptors Hemocyanins biology.protein Female Immunization Antibody Peptides Haptens Keyhole limpet hemocyanin |
| Zdroj: | Monoclonal Antibodies in Immunodiagnosis and Immunotherapy. 34:36-43 |
| ISSN: | 2167-9436 |
| DOI: | 10.1089/mab.2014.0077 |
| Popis: | The gene that codes for the CD44 family members consists of 20 exons, nine of which encode the standard form of the molecule. The other exons can be inserted in various combinations into the membrane proximal region of the extracellular domain of the protein, giving rise to variant isoforms (CD44v). CD44 variants, especially the CD44v6, have been reported to regulate tumor invasion, progression, and metastasis of carcinomas. Producing a high affinity monoclonal antibody against human CD44v6 provides a powerful tool to monitor and trace CD44v6 function in different biological fluids. In this study, a synthetic peptide from CD44v6 was conjugated to keyhole limpet hemocyanin (KLH) and injected into BALB/c mice. Splenocytes from the immunized mice were fused with murine SP2/0 myeloma cells followed by selection of antibody producing hybridoma cells. After screening of hybridoma colonies by ELISA, high affinity antibodies were selected and purified by affinity chromatography. Western blot, immunocytochemistry, and flow cytometry experiments were used to characterize the antibodies. Six stable hybridoma cell lines, designated as 1H1, 1H2, 2A12, 2G11, 3H3, and 3H7, were obtained. Flow cytometry and immunocytochemistry results showed that the new monoclonal antibodies recognized CD44v6 on the cell surface. This novel panel of anti-CD44v6 antibodies has the potential for investigating the role of CD44v6 in cancer pathogenesis. |
| Databáze: | OpenAIRE |
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