Browsing by keyword "Phage display"
Now showing items 1-2 of 2
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A new TAG-72 cancer marker peptide identified by phage displayRadiolabeled peptides as markers of cancer targets have demonstrated their value in diagnostic imaging and radiotherapy. The 16 mer f88-4/Cys6 phage display library was applied to affinity purified TAG-72 and three consensus peptides were identified: VHHSCTKLTHCCQNWH (A2-13), GGVSCMQTSPVCENNL (A2-6) and TKRDCSAQNYGCQKAI (A2-11). The A2-13 and A2-6 phages showed the highest percent binding to LS-174T cells by flow cytometry and were 3-fold higher than a control phage, while fluorescence microscopy showed that both A2-6 and A2-13 phages bound to the LS-174T cell membrane. However, only the A2-6 phage demonstrated specificity by low binding to the TAG-72 negative cell HT-29. Furthermore, the synthesized free A2-6 peptide demonstrated specific binding to LS-174T cells by flow cytometry and by immunohistochemical staining of xenograft tumor compared to normal colon. These data indicate that the A2-6 peptide is specific for the TAG-72 cancer target.
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Isolation of a monoclonal antibody from a phage display library binding the rhesus macaque MHC class I allomorph Mamu-A1*001Monoclonal antibodies that bind to human leukocyte antigen (HLA) are useful tools for HLA-typing, tracking donor-recipient chimerisms after bone marrow transplants, and characterizing specific major histocompatibility complexes (MHC) on cell surfaces. Unfortunately, equivalent reagents are not available for rhesus macaques, which are commonly used animal as models in organ transplant and infectious disease research. To address this deficiency, we isolated an antibody that recognizes the common Indian rhesus macaque MHC class I molecule, Mamu-A1*001. We induced Mamu-A1*001-binding antibodies by alloimmunizing a female Mamu-A1*001-negative rhesus macaque with peripheral blood mononuclear cells (PBMC) from a male Mamu-A1*001-positive donor. A Fab phage display library was constructed with PBMC from the alloimmunized macaque and panned to isolate an antibody that binds to Mamu-A1*001 but not to other common rhesus macaque MHC class I molecules. The isolated antibody distinguishes PBMC from Mamu-A1*001-positive and -negative macaques. Additionally, the Mamu-A1*001-specific antibody binds the cynomolgus macaque MHC class I ortholog Mafa-A1*001:01 but not variants Mafa-A1*001:02/03, indicating a high degree of binding specificity. The Mamu-A1*001-specific antibody will be useful for identifying Mamu-A1*001-positive rhesus macaques, for detecting Mamu-A1*001-positive cells in populations of Mamu-A1*001-negative cells, and for examining disease processes that alter expression of Mamu-A1*001 on cell surfaces. Moreover, the alloimmunization process we describe will be useful for isolating additional MHC allomorph-specific monoclonal antibodies or antibodies against other polymorphic host proteins which are difficult to isolate with traditional technologies.
