Structural characterization of the epitope recognized by the new anti‐Fy6 monoclonal antibody NaM185‐2C3

Abstract

The epitope recognized by a new anti-Fy6 monoclonal antibody (MoAb) (clone name: NaM185-2C3) was characterized using peptides synthesized on pins (Epitope scanning kit). The clone was obtained from splenocytes of mice immunized with CHO cells expressing the recombinant Duffy glycoprotein. NaM185-2C3 recognized a linear epitope, the essential portion of which was pentapeptide Phe-Glu-Asp-Val-Trp comprising amino acid residues 22-26 of the main (336aa) isoform of the Duffy antigen. All the amino acid residues of the epitope, except Asp, were essential for the antibody-binding, because they could not be replaced by any or most other amino acid residues. The Asp residue could be replaced by most other amino acid residues and its replacement by some amino acid residues gave a distinct increase in the antibody-binding. The MoAb NaM185-2C3, similarly as other anti-Fy6 antibodies, inhibits interleukin (IL)-8-binding to the Duffy antigen. A part of the results was presented at ISBT meeting (Blanchard et al., 1998, Vox Sanguinis, 74, S1, Abstract no. 71).

Enhanced Expression of Duffy Antigen in the Lungs During Suppurative Pneumonia

“…(Tournamille et al 1998;Yazdanbakhsh et al 2000). Because the Fy x polymorphism results from a point mutation within the coding region of the Duffy gene, the nonerythroid cells should also show reduced expression and chemokine binding (Olsson et al 1998;Parasol et al 1998;Tournamille et al 1998;Wasniowska et al 2002).…”

“…Endogeous peroxidases and biotin were blocked using 3% H 2 O 2 in methanol and an avidinbiotin blocking kit (Vector Labs). The specificity of MAb ␣ Fy6 (clone 2C3) used in this study was determined by immunoblotting, and epitope mapping showed that it binds to the epitope 24 FEDVW 28 located on the amino-terminal extracellular domain of the Duffy glycoprotein (Segerer et al 2000;Wasniowska et al 2002). An irrelevant monoclonal IgG 1 isotype antibody (Dako Diagnostics; Glostrup, Denmark) was used as the control primary antibody.…”

Lymphatic Precollectors Contain a Novel, Specialized Subpopulation of Podoplaninlow, CCL27-Expressing Lymphatic Endothelial Cells

“…555445; BD Biosciences Pharmingen, Franklin Lakes, NJ), RPE-Cy5.1-conjugated mouse monoclonal anti-CD45 (catalog no. PM IM2653; Beckman Coulter, Inc., Fullerton, CA), mouse clone 2C3 anti-Duffy blood group antigen receptor for chemokines (DARC)/Fy (kindly provided by Dr. Ives Colin; INSERM, Institute National de Transfusion Sanguine, Paris 5,6 ), mouse monoclonal anti-CCL27 (catalog no. MAB3761; R&D Systems, Minneapolis, MN), neutralizing rat anti-CCL27 antibody (clone 68623; catalog no.…”

Strict tropism for CD71+/CD234+ human reticulocytes limits the zoonotic potential of Plasmodium cynomolgi

“…Cultures were incubated at 37°C for at least 24 hours, which allowed the merozoites to reinvade fresh RBCs. Blockade of RBC invasion was assayed by using different antibodies and a chemokine against human DARC: anti-FY6 2C3 25 at 20 mg/mL, antiFyB (Abcam) at 25 mg/mL, and the chemokine interleukin-8 (CXCL-8/IL-8) (Peprotech Asia) at 1000 nM. The overall experimental scheme is shown in Figure 1.…”