Production of Recombinant 14-3-3 Protein and Determination of Its Immunogenicity for Application in Serodiagnosis of Strongyloidiasis Publisher Pubmed



Masoori L¹ ; Falak R^{2, 3} ; Mokhtarian K⁴ ; Bandehpour M^{5, 6} ; Razmjou E¹ ; Jalallou N⁷ ; Jafarian F⁸ ; Akhlaghi L¹ ; Meamar AR¹ Show Affiliations
Source: Transactions of the Royal Society of Tropical Medicine and Hygiene Published:2019

Abstract

Background: Strongyloides stercoralis is the fourth most important intestinal nematode worldwide. The parasite load and larvae count are often low, thus conventional methods are not sufficiently sensitive to detect the infection. In this study we developed an immunoglobulin G-based enzyme-linked immunosorbent assay (ELISA) method to detect antibodies against S. stercoralis 14-3-3 protein in patients' sera. Methods: S. stercoralis RNA was extracted and following complementary DNA synthesis, the 708-bp fragment of 14-3-3 protein was amplified by polymerase chain reaction and cloned into the pET28a+ expression vector. The 30-kDa recombinant 14-3-3 protein was expressed in Escherichia coli BL21 (DE3) cells and purified by affinity chromatography. Finally, its immunoreactivity was assessed by indirect ELISA and western blotting. Results: The S. stercoralis 14-3-3 gene was successfully amplified and cloned into an expression vector. The 30-kDa recombinant protein was purified by affinity chromatography. An ELISA developed in-house detected infected patients' sera with 96% sensitivity. Conclusions: We concluded that the recombinant 14-3-3 protein has enough sensitivity and specificity for detection of strongyloidiasis in human sera and could be applied for serodiagnosis. © 2019 The Author(s).