Antigen capture enzyme-linked immunosorbent assay for specific detection of Reston Ebola virus nucleoprotein

Abstract

Antigen capture enzyme-linked immunosorbent assay (ELISA) is one of the most useful methods to detect Ebola virus rapidly. We previously developed an antigen capture ELISA using a monoclonal antibody (MAb), 3-3D, which reacted not only to the nucleoprotein (NP) of Zaire Ebola virus (EBO-Z) but also to the NPs of Sudan (EBO-S) and Reston Ebola (EBO-R) viruses. In this study, we developed antigen capture ELISAs using two novel MAbs, Res2-6C8 and Res2-1D8, specific to the NP of EBO-R. Res2-6C8 and Res2-1D8 recognized epitopes consisting of 4 and 8 amino acid residues, respectively, near the C-terminal region of the EBO-R NP. The antigen capture ELISAs using these two MAbs detected the EBO-R NP in the tissues from EBO-R-infected cynomolgus macaques. The antigen capture ELISAs using Res2-6C8 and Res2-1D8 are useful for the rapid detection of the NP in EBO-R-infected cynomolgus macaques.

FIG. 1.

IFA using HeLa cells expressing rNP of EBO-R (A to C) or EBO-Z (D-F) and using Vero cells infected with EBO-S (G to I). The reactivities of Res2-6C8 (A, D, and G), Res2-1D8 (B, E, and H), and 3-3D (C, F, and I) to NP of EBO-R, EBO-Z, and EBO-S are shown. Res2-6C8 and Res2-1D8 reacted to EBO-R NP, but not to the NPs of EBO-Z and EBO-S. The MAb 3-3D reacts to the NPs of all three Ebola virus species.

FIG. 2.

Reactivities of Res2-6C8 and Res2-1D8 to the truncated EBO-R rNPs in IgG ELISA. Res2-6C8 recognizes the amino acid residues between aa 636 and 639 (sequence, DPDI). Res2-1D8 recognizes the amino acid residues between aa 636 and 643 (sequence, DPDIGQSK). The mean OD value ± SD in the IgG ELISA is shown.

FIG. 3.

Reactivity of MAbs Res2-6C8 (□), Res2-1D8 (▵), and 3-3D (♦) to the rNPs of EBO-R and EBO-Z in the antigen capture ELISA. His-EBO-R-NP (A) and His-EBO-Z-NP (B) were used as antigens. The mean ± SD of four assays is shown.