Development of an enzyme-linked immunosorbent assay using recombinant protein antigen for the diagnosis of Chikungunya virus

Flávia Fonseca Bagno,Lara Carvalho Godoi,Natalia Salazar,Glauco de Carvalho Pereira,Maria Marta Figueiredo,Flávio Guimarães da Fonseca

doi:10.1016/j.dib.2019.104015

Flávia Fonseca Bagno, Lara Carvalho Godoi + Show 4 more

Open Access

https://doi.org/10.1016/j.dib.2019.104015

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Abstract

We describe here the development of an in-house enzyme linked immunosorbent assay (ELISA) for the diagnostic of Chikungunya virus (CHIKV) infections using a recombinant protein from CHIKV. The recombinant protein gene was designed based on 154 sequences and we used computational methods to predict its structure and antigenic potential. To confirm predictions, the gene coding for the recombinant CHIKV protein (rCHIKVp) was synthetized and expressed in prokaryotic system. Subsequently, the protein was purified by affinity chromatography and used as antigen in an indirect ELISA. We present data regarding the optimization of the recombinant antigen production and preparation of the ELISA to detect IgG against CHIKV in human sera.

Highlights

We describe here the development of an in-house enzyme linked immunosorbent assay (ELISA) for the diagnostic of Chikungunya virus (CHIKV) infections using a recombinant protein from CHIKV
We present data regarding the optimization of the recombinant antigen production and preparation of the ELISA to detect immunoglobulin G (IgG) against CHIKV in human sera
The ELISA's recombinant antigen - the E2 protein from Chikungunya virus - was expressed in E. coli features transformed cells and purified by affinity chromatography

Summary

Design of the antigen gene

CHIKV genome sequences deposited at GenBank were aligned using MEGA7 software [2]. The selection criteria were: complete annotation of the genome and absence of indefinite nucleotides in the sequence. Sequences belonged to the Asian genotype (n 1⁄4 112) and East-Central South African (ECSA). A consensus sequence was generated for each group (Group 1: Brazilian samples, Group 2: Asian genotype samples and Group 3: ECSA genotype samples). These consensual sequences were, compared to each other to generate a unique nucleotide sequence. From these predictions, a cut-off point for the generation of the protein without its transmembrane domain was determined. The nucleotide sequence of the truncated protein-coding gene was commercially synthesized and subcloned into the pET-21 expression vector, which included a histidine tag to the construction

In silico analysis

Recombinant protein production

In house Anti-Chikungunya virus ELISA