Open AccessEnhancing the detection of Toxoplasma gondii via an anti-SAG1 scFv-alkaline phosphatase immunoconjugate
Author(s) -
Emna Hannachi,
Aïda Bouratbine,
Mohamed Mousli
Publication year - 2019
Publication title -
biotechnology reports
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.932
H-Index - 36
ISSN - 2215-017X
DOI - 10.1016/j.btre.2019.e00360
Subject(s) - immunoconjugate , toxoplasma gondii , immunoassay , recombinant dna , immunofluorescence , conjugate , alkaline phosphatase , microbiology and biotechnology , chemistry , antibody , biology , monoclonal antibody , biochemistry , enzyme , gene , immunology , mathematical analysis , mathematics
The purpose of this study was to implement a fluorometric method for enhancing the detection sensitivity of in biological fluids. To address this challenge, we designed and produced a recombinant immunoconjugate tool based on a single-chain antibody fragment anti- SAG1 antigen (scFvSG15) genetically fused to the bacterial alkaline phosphatase (AP) using 4-methyl-umbelliferyl-phosphate as fluorogenic substrate. The anti-SAG1 scFv-AP conjugate was fully bifunctional and was used successfully in different assays including immunoblot, fluorometric ELISA and direct immunofluorescence. The fluorometric immunoassay afforded an extremely low detection limit (1 tachyzoite/well), which was in agreement with the real-time PCR control test. The immunofluorescence imaging has provided captivating visual evidence of detection. These results strongly suggest that the recombinant anti-SAG1-AP conjugate generated here might serve as useful and highly sensitive immunoassay probe to direct detect in a one-step procedure, opening up new perspectives for diagnosis of toxoplasmosis.
Accelerating Research
Robert Robinson Avenue,
Oxford Science Park, Oxford
OX4 4GP, United Kingdom
Address
John Eccles HouseRobert Robinson Avenue,
Oxford Science Park, Oxford
OX4 4GP, United Kingdom