Efficient generation of recombinant anti-HER2 scFv with high yield and purity using a simple method.
Biotechnol J
; 19(7): e2300745, 2024 Jul.
Article
en En
| MEDLINE
| ID: mdl-39014926
ABSTRACT
We developed a method to produce a soluble form of a single-chain fragment variable (scFv) targeting human epithelial growth factor receptor 2 (HER2) in Escherichia coli. By optimizing the orientations of the variable heavy (VH) and variable light (VL) domains and the His-tag, we identified the HL-His type antibody with the highest HER2-binding activity. Purification of HL-His yielded 40.7 mg from a 1 L culture, achieving >99% purity. The limit of detection was determined to be 2.9 ng, demonstrating high production yield, purity, and sensitivity. Moreover, we successfully labeled HER2+ cell lines with fluorescent dye-conjugated scFv, resulting in a significantly higher observed signal-to-background ratio, compared to that of HER2- cell lines. This highlights the potential of these fluorescent scFvs as valuable probes for HER2+ breast cancer diagnostics. Notably, the process for the complete scFv production was streamlined and required only 4-5 days. Additionally, the product maintained its activity after freeze storage, allowing for large-scale production and a wide range of practical applications.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Proteínas Recombinantes
/
Receptor ErbB-2
/
Escherichia coli
/
Anticuerpos de Cadena Única
Límite:
Humans
Idioma:
En
Revista:
Biotechnol J
Asunto de la revista:
BIOTECNOLOGIA
Año:
2024
Tipo del documento:
Article