Development of a New Highly Selective Monoclonal Antibody against Preferentially Expressed Antigen in Melanoma (PRAME) and Identification of the Target Epitope by Bio-Layer Interferometry
Development of a New Highly Selective Monoclonal Antibody against Preferentially Expressed Antigen in Melanoma (PRAME) and Identification of the Target Epitope by Bio-Layer Interferometry(303 views) Sivaccumar JP, Leonardi A, Iaccarino E, Corvino G, Sanguigno L, Chambery A, Russo R, Valletta M, Latino D, Capasso D, Doti N, Ruvo M, Sandomenico A
Int J Mol Sc (ISSN: 1422-0067linking, 1422-0067electronic, 1661-6596), 2021 Mar 20; 22(6): N/D-N/D.
Istituto di Biostrutture e Bioimmagini, CNR, 80134 Napoli, Italy.
Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Università degli Studi di Napoli Federico II, 80142 Napoli, Italy.
Dipartimento di Scienze e Tecnologie Ambientali, Biologiche e Farmaceutiche (DISTABIF), Università L. Vanvitelli, 80100 Caserta, Italy.
Centro di Servizio di Ateneo per le Scienze e Tecnologie per la Vita (CESTEV), Università di Napoli Federico II, 80145 Napoli, Italy.
References: Not available.
Development of a New Highly Selective Monoclonal Antibody against Preferentially Expressed Antigen in Melanoma (PRAME) and Identification of the Target Epitope by Bio-Layer Interferometry
BACKGROUND: Monoclonal antibodies (mAbs) against cancer biomarkers are key reagents in diagnosis and therapy. One such relevant biomarker is a preferentially expressed antigen in melanoma (PRAME) that is selectively expressed in many tumors. Knowing mAb's epitope is of utmost importance for understanding the potential activity and therapeutic prospective of the reagents. METHODS: We generated a mAb against PRAME immunizing mice with PRAME fragment 161-415; the affinity of the antibody for the protein was evaluated by ELISA and SPR, and its ability to detect the protein in cells was probed by cytofluorimetry and Western blotting experiments. The antibody epitope was identified immobilizing the mAb on bio-layer interferometry (BLI) sensor chip, capturing protein fragments obtained following trypsin digestion and performing mass spectrometry analyses. RESULTS: A mAb against PRAME with an affinity of 35 pM was obtained and characterized. Its epitope on PRAME was localized on residues 202-212, taking advantage of the low volumes and lack of fluidics underlying the BLI settings. CONCLUSIONS: The new anti-PRAME mAb recognizes the folded protein on the surface of cell membranes suggesting that the antibody's epitope is well exposed. BLI sensor chips can be used to identify antibody epitopes.
Development of a New Highly Selective Monoclonal Antibody against Preferentially Expressed Antigen in Melanoma (PRAME) and Identification of the Target Epitope by Bio-Layer Interferometry
Development of a New Highly Selective Monoclonal Antibody against Preferentially Expressed Antigen in Melanoma (PRAME) and Identification of the Target Epitope by Bio-Layer Interferometry
Bogdanovich S, Kim Y, Mizutani T, Yasuma R, Tudisco L, Cicatiello V, Bastos-carvalho A, Kerur N, Hirano Y, Baffi JZ, Tarallo V, Li S, Yasuma T, Arpitha P, Fowler BJ, Wright CB, Apicella I, Greco A, Brunetti A, Ruvo M, Sandomenico A, Nozaki M, Ijima R, Kaneko H, Ogura Y, Terasaki H, Ambati BK, Leusen JH, Langdon WY, Clark MR, Armour KL, Bruhns P, Verbeek JS, Gelfand BD, De Falco S, Ambati J * Human IgG1 antibodies suppress angiogenesis in a target-independent manner(1007 views) Signal Transduct Target Ther (ISSN: 2059-3635print), 2016; 1: N/D-N/D. Impact Factor:0 ViewExport to BibTeXExport to EndNote