Detection of Amino-terminal Extracellular Domain of Somatostatin Receptor 2 by Specific Monoclonal Antibodies and Quantification of Receptor Density in Medulloblastoma
| Autor: | Roger E. McLendon, Ujendra Kumar, Darell D. Bigner, Carol J. Wikstrand, Michael R. Zalutsky, Chien-Tsun Kuan |
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| Rok vydání: | 2009 |
| Předmět: |
medicine.drug_class
media_common.quotation_subject Blotting Western Immunology Enzyme-Linked Immunosorbent Assay Biology Monoclonal antibody Epitope Cell Line Tumor medicine Humans Immunology and Allergy Somatostatin receptor 2 Amino Acid Sequence Receptors Somatostatin Receptor Internalization DNA Primers media_common Reverse Transcriptase Polymerase Chain Reaction Antibodies Monoclonal Original Articles Flow Cytometry Ligand (biochemistry) Immunohistochemistry Molecular biology Protein Structure Tertiary Cell culture biology.protein Immunotherapy Antibody Medulloblastoma |
| Zdroj: | Hybridoma. 28:389-403 |
| ISSN: | 1557-8348 1554-0014 |
| Popis: | Somatostatin receptor 2 (SSTR2) is expressed by most medulloblastomas (MEDs). We isolated monoclonal antibodies (MAbs) to the 12-mer (33)QTEPYYDLTSNA(44), which resides in the extracellular domain of the SSTR2 amino terminus, screened the peptide-bound MAbs by fluorescence microassay on D341 and D283 MED cells, and demonstrated homogeneous cell-surface binding, indicating that all cells expressed cell surface-detectable epitopes. Five radiolabeled MAbs were tested for immunoreactive fraction (IRF), affinity (KA) (Scatchard analysis vs. D341 MED cells), and internalization by MED cells. One IgG(3) MAb exhibited a 50-100% IRF, but low KA. Four IgG(2a) MAbs had 46-94% IRFs and modest KAs versus intact cells (0.21-1.2 x 10(8) M(-1)). Following binding of radiolabeled MAbs to D341 MED at 4 degrees C, no significant internalization was observed, which is consistent with results obtained in the absence of ligand. However, all MAbs exhibited long-term association with the cells; binding at 37 degrees C after 2 h was 65-66%, and after 24 h, 52-64%. In tests with MAbs C10 and H5, the number of cell surface receptors per cell, estimated by Scatchard and quantitative FACS analyses, was 3.9 x 10(4) for the "glial" phenotype DAOY MED cell line and 0.6-8.8 x 10(5) for four neuronal phenotype MED cell lines. Our results indicate a potential immunotherapeutic application for these MAbs. |
| Databáze: | OpenAIRE |
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