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Preparation and application of a specific single-chain variable fragment against artemether
Fang Lu1, Fa Zhang1, Jingqi Qian2
1School of Biotechnology and Health Sciences, Wuyi University, 529020 Jiangmen, Guangdong, China.
Journal of Pharmaceutical and Biomedical Analysis
|September 1, 2022
Summary
Researchers developed a genetically engineered antibody fragment (scFv) specific to artemether. This scFv enabled a sensitive indirect competitive ELISA for accurately quantifying artemether in drugs, consistent with UPLC analysis.
Area of Science:
- Biotechnology
- Immunology
- Analytical Chemistry
Background:
- Artemether is a key component in artemisinin-based combination therapies, crucial for malaria treatment.
- Monoclonal antibodies offer high specificity for drug detection, but their application can be limited.
- Developing recombinant antibody fragments like single-chain variable fragments (scFv) can enhance utility and production.
Purpose of the Study:
- To clone and express an artemether-specific single-chain variable fragment (scFv).
- To develop a sensitive and specific indirect competitive enzyme-linked immunosorbent assay (icELISA) for artemether detection.
- To validate the developed icELISA by comparing its results with ultra-performance liquid chromatography (UPLC).
Main Methods:
- Cloning of VH and VL genes from an artemether-specific monoclonal antibody (mAb 2G12E1).
- Construction of scFv into various prokaryotic expression vectors, with successful soluble expression achieved using pMAL-p5x.
- Development of an indirect competitive ELISA using the anti-artemether scFv.
Main Results:
- Soluble expression of anti-artemether scFv was achieved using the pMAL-p5x vector, maintaining sensitivity and specificity.
- The developed icELISA demonstrated a low IC50 value of 4.33 ng/mL and a working range of 1.05-22.65 ng/mL.
- Artemether quantification using icELISA showed high consistency with results obtained via UPLC analysis.
Conclusions:
- The anti-artemether scFv is a valuable tool for artemether detection and monitoring.
- The developed icELISA provides a reliable method for quantifying artemether content in pharmaceutical preparations.
- This genetically engineered antibody fragment has potential applications in drug quality control and mechanistic studies.
