Abstract
Phage display is a powerful technology that selects specific proteins or peptides to a target. We have used Phage Display to select scFv (single-chain variable fragment) clones from a combinatorial library against total proteins of Strongyloides venezuelensis. After scFv characterization, further analysis demonstrated that this recombinant fragment of antibody was able to bind to an S. venezuelensis antigenic fraction of ~65 kDa, present in the body periphery and digestive system of infective larvae (L3), as demonstrated by immunofluorescence. Mass spectrometry results followed by bioinformatics analysis showed that this antigenic fraction was a heat shock protein 60 (HSP60) of Strongyloides sp. The selected scFv was applied in serodiagnosis by immune complexes detection in serum samples from individuals with strongyloidiasis using a sandwich enzyme-linked immunosorbent assay (ELISA), showing sensitivity of 97.5% (86.84–99.94), specificity of 98.81 (93.54–99.97), positive likelihood ratio of 81.60 and an area under the curve of 0.9993 (0.9973–1.000). Our study provided a novel monoclonal scFv antibody fragment which specifically bound to HSP60 of Strongyloides sp. and was applied in the development of an innovative serodiagnosis method for the human strongyloidiasis.
Highlights
Phage display is a powerful technology that selects specific proteins or peptides to a target
The engineering of novel functional combinatorial antibodies expressed on the surface of bacteriophages, such as scFvs, with high specificity for antigens coupled with a phage display strategy is the most promising technology for improved diagnostics and therapeutics of many pathologies[10,11]
This is due to their small size, great reactivity and specificity, and easiness to maintain and express in bacteria, which surpass the hybridoma technology in all aspects
Summary
Phage display is a powerful technology that selects specific proteins or peptides to a target. We have used Phage Display to select scFv (single-chain variable fragment) clones from a combinatorial library against total proteins of Strongyloides venezuelensis. The major problem for the serodiagnosis of human strongyloidiasis is caused the difficulty in obtaining larvae of Strongyloides stercoralis for antigen preparation. Because of this difficulty, heterologous antigens from Strongyloides venezuelensis have been used for convenience, due to similarities in transcripts that have key roles in the host-parasite interaction, as well as important molecules for diagnosis such as excretory/secretory proteins. Phage display is a method which provides the selection of peptides, antibodies or single-chain variable fragment (scFv), expressed on bacteriophages, by standard methods with shorter time of production and important applicability in diagnosis due high specificity of the selected molecule to the target[6,7,8,9]. The structure of the selected scFv and its binding was characterized by www.nature.com/scientificreports/
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