Screening for single-chain variable fragment antibodies against multiple Cry1 toxins from an immunized mouse phage display antibody library

Applied Microbiology and Biotechnology
Sa DongXianjin Liu

Abstract

Single-chain variable fragment (scFv) is a kind of antibody that possess only one chain of the complete antibody while maintaining the antigen-specific binding abilities and can be expressed in prokaryotic system. In this study, scFvs against Cry1 toxins were screened out from an immunized mouse phage displayed antibody library, which was successfully constructed with capacity of 6.25 × 107 CFU/mL. Using the mixed and alternative antigen coating strategy and after four rounds of affinity screening, seven positive phage-scFvs against Cry1 toxins were selected and characterized. Among them, clone scFv-3H9 (MG214869) showing relative stable and high binding abilities to six Cry1 toxins was selected for expression and purification. SDS-PAGE indicated that the scFv-3H9 fragments approximately 27 kDa were successfully expressed in Escherichia coli HB2151 strain. The purified scFv-3H9 was used to establish the double antibody sandwich enzyme-linked immunosorbent assay method (DAS-ELISA) for detecting six Cry1 toxins, of which the lowest detectable limits (LOD) and the lowest quantitative limits (LOQ) were 3.14-11.07 and 8.22-39.44 ng mL-1, respectively, with the correlation coefficient higher than 0.997. The average recoveries of Cry1...Continue Reading

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Citations

May 22, 2019·Biotechnology and Bioengineering·Marsela JorgolliLes P Miranda
May 19, 2021·International Journal of Biological Macromolecules·Sa DongJianguo Feng
Jul 27, 2021·Frontiers in Cellular and Infection Microbiology·Kristian Daniel Ralph RothMichael Hust
Jul 29, 2021·Journal of Agricultural and Food Chemistry·Aroosha FaheemYonggang Hu

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Datasets Mentioned

BETA
MG214869

Methods Mentioned

BETA
enzyme-linked immunosorbent assay
ELISA
phage display
PCR
electrophoresis
glycosylation

Software Mentioned

SPSS

Related Concepts

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