| Autor: |
Gaffar, Shabarni, Nurbayanti, Siti Hesti, Hartati, Yeni Wahyuni, Novianti, Mia Tria, Novitriani, Korry, Ishmayana, Safri, Yusuf, Muhammad, Subroto, Toto |
| Předmět: |
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| Zdroj: |
Journal of Immunoassay & Immunochemistry; 2024, Vol. 45 Issue 4, p307-324, 18p |
| Abstrakt: |
Single Chain Variable Fragment (scFv), a small fragment of antibody can be used to substitute the monoclonal antibody for diagnostic purposes. Production of scFv in Escherichia coli host has been a challenge due to the potential miss-folding and formation of inclusion bodies. This study aimed to express anti-CHIKV E2 scFv which previously designed specifically for Asian strains by co-expression of three chaperones that play a role in increasing protein solubility; GroEL, GroES, and Trigger Factor. The scFv and chaperones were expressed in Origami B E. coli host under the control of the T7 promoter, and purified using a Ni-NTA column. Functional assay of anti-CHIKV-E2 scFv was examined by electrochemical immunosensor using gold modified Screen Printed Carbon Electrode (SPCE), and characterized by differential pulses voltammetry (DPV) using K3[Fe(CN)6] redox system and scanning microscope electron (SEM). The experimental condition was optimized using the Box-Behnken design. The results showed that co-expression of chaperone increased the soluble scFv yield from 54.405 μg/mL to 220.097 µg/mL (~5×). Furthermore, scFv can be used to detect CHIKV-E2 in immunosensor electrochemistry with a detection limit of 0.74048 ng/mL and a quantification limit of 2,24388 ng/mL. Thus, the scFv-anti-CHIKV-E2 can be applied as a bioreceptor in another immunoassay method. [ABSTRACT FROM AUTHOR] |
| Databáze: |
Complementary Index |
| Externí odkaz: |
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