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Design and expression of polytopic construct of cathepsin-L1, SAP-2 and FhTP16.5 proteins of Fasciola hepatica

Published online by Cambridge University Press:  04 March 2020

S. Aghamolaei ,
B. Kazemi ,
M. Bandehpour ,
M.M. Ranjbar ,
S. Rouhani ,
A. Javadi Mamaghani  and
S.J.S. Tabaei Open the ORCID record for S.J.S. Tabaei [Opens in a new window]
S. Aghamolaei
Affiliation:
Department of Parasitology and Mycology, School of Medicine, Student Research Committee, Shahid Beheshti University of Medical Sciences, Tehran, Iran
B. Kazemi
Affiliation:
Cellular & Molecular Biology Research Center, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
M. Bandehpour
Affiliation:
Cellular & Molecular Biology Research Center, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
M.M. Ranjbar
Affiliation:
Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran
S. Rouhani
Affiliation:
Department of Parasitology and Mycology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
A. Javadi Mamaghani
Affiliation:
Department of Parasitology and Mycology, School of Medicine, Student Research Committee, Shahid Beheshti University of Medical Sciences, Tehran, Iran
S.J.S. Tabaei*
Affiliation:
Department of Parasitology and Mycology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
*
Author for correspondence: S.J.S. Tabaei, E-mail: [email protected]
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Abstract

The enzyme-linked immunosorbent assay (ELISA) technique can play an important role in the early detection of fascioliasis. However, they have some diagnostic limitations, including cross-reaction with other helminths. It seems that the combination of recombinant parasite proteins as antigen can reduce these problems. Hence, the present study was aimed to design and confirm the antigenic recombinant multi-epitope (rMEP) construct of three protein epitopes (linear and conformational B-cell epitopes) of the parasite using immunoinformatic tools. For this purpose, the tertiary structures of Fasciola hepatica cathepsin-L1, saposin-like protein 2 and 16.5-kDa tegument-associated protein were predicted using the I-TASSER server. Validation of the modelled structures was performed by Ramachandran plots. The antigenic epitopes of the proteins were achieved by analysing the features of the IEDB server. The synthesized gene was cloned into the pET-22b (+) expression vector and transformed into the Escherichia coli BL21. Sodium dodecyl sulfate polyacrylamide gel electrophoresis was used to verify and analyse the expression of the rMEP protein. Western blotting was utilized to confirm rMEP protein immunogenicity in two forms, one using an anti-His tag antibody and the other with human pooled sera samples (fascioliasis, non-fascioliasis and negative control sera). Our results demonstrated that the rMEP designed for the three proteins of F. hepatica was highly antigenic, and immune-detection techniques confirmed the antigen specificity. In conclusion, the presented antigenic multi-epitope may be very helpful to develop serodiagnostic kits such as indirect ELISA to evaluate the proper diagnosis of fascioliasis in humans and ruminants.

Keywords

FascioliasisFasciolaepitopescathepsin-L1SAP-2FhTP16.5recombinant proteinsmulti-epitopeimmunoinformatic
Type
Research Paper
Information
Journal of Helminthology , Volume 94 , 2020 , e134
Copyright
Copyright © Cambridge University Press 2020

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