Development of a sensitive direct competitive chemiluminescent enzyme immunoassay for gentamicin based on the construction of a specific single-chain variable fragment-alkaline phosphatase fusion protein.
Microchem J
; 1972024 Feb.
Article
em En
| MEDLINE
| ID: mdl-38283367
ABSTRACT
A sensitive chemiluminescent enzyme immunoassay (CLEIA) was established for the determination of gentamicin (GEN) residue levels in animal tissue. This assay is based on a fusion protein of single-chain variable fragment (scFv) and alkaline phosphatase (AP). Initially, VL and VH derived from anti-gentamicin monoclonal antibody were linked by a short peptide to construct a scFv. Subsequently, the constructed scFv sequence was accessed into the pLIP6/GN vector, and a soluble scFv-AP fusion protein was generated. The scFv-AP fusion protein was used to develop a direct competitive CLEIA (dcCLEIA) for the determination of gentamicin. In the dcCLEIA, the half inhibitory concentration (IC50) and limit of detection (LOD) were 1.073 ng/mL and 0.380 ng/mL, respectively. The average recoveries of gentamicin spiked in animal tissue samples ranged from 78% to 96%. These results showed a strong correlation with ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). The above results suggest that the anti-GEN scFv-AP fusion protein is suitable for detecting gentamicin residues in edible animal tissues.
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1
Coleções:
01-internacional
Base de dados:
MEDLINE
Tipo de estudo:
Diagnostic_studies
Idioma:
En
Revista:
Microchem J
Ano de publicação:
2024
Tipo de documento:
Article