Affinity maturation of single-chain variable fragment specific for aflatoxin B1 using yeast surface display. (1st December 2015)

• Record Type: Journal Article
• Authors: Min, Won-Ki; Kim, Sung-Gun; Seo, Jin-Ho
• Is Part Of: Food chemistry. Volume 188(2015)
• Page Start: 604
• Page End: 611
• Publication Date: 2015-12-01
• Subjects: Single-chain variable fragment -- Aflatoxin B1 -- Yeast surface display -- Fluorescence-activated cell sorting; Food -- Analysis -- Periodicals; Food -- Composition -- Periodicals; 664
• DOI: 10.1016/j.foodchem.2015.04.117
• Languages: English
• ISSNs: 0308-8146

Abstract

Highlights: Yeast scFv library was established for FACS screening. 5 scFvs with improved affinity against AFB1 were isolated via 4 rounds of screening. The best scFv with 9-fold improvement in affinity represents high specificity to AFB1 . Abstract: As aflatoxin B1 is one of the most toxic mycotoxins, it is important to detect and to quantify aflatoxin B1 accurately by immunological methods. To enhance aflatoxin B1 -binding affinity of the single-chain variable fragment, yeast surface display technique combined with fluorescence-activated cell sorting was applied. A randomly mutated scFv library was subjected to 4 rounds of fluorescence-activated cell sorting, resulting in isolation of 5 scFv variants showing an affinity improvement compared to the parental wild type scFv. The best scFv with a 9-fold improvement in affinity for aflatoxin B1 exhibited similar specificity to the monoclonal antibody. Most of the mutations in scFv-M37 were located outside of the canonical antigen-contact loops, suggesting that its affinity improvement might be driven by an allosteric effect inducing scFv-M37 to form a more favorable binding pocket for aflatoxin B1 than the wild type scFv.