Generation of panels of anti-idiotypic single-chain variable fragments mimicking Cry2Aa toxin using the chain shuffling technique

Abstract

Here, we firstly reported to generate panels of anti-idiotypic singlechain variable fragments (scFvs) of Bacillus thuringiensis Cry2Aa toxin by chain shuffling method. Light-and heavy-chain-shuffled libraries based on two parental clones were constructed. The signal of the light-chain-shuffled library was 4.2-fold higher than the heavy one. Solely after one round panning, eight mutants were rapidly isolated from the chain-shuffled libraries. The apparent affinity of the best clones was 2.65 × 10 6 M −1 , a 2.59-fold improvement over the parental clone. The binding abilities between anti-idiotypic scFvs with brush border membrane vesicle of Plutella xylostella larvae were also determined. This work demonstrated that chain shuffling could rapidly generate panels of antibodies with similar binding abilities but of the distinct idiotypic composition. The serials of mutants can be used to map the binding epitope of the receptors in target insects instead of Cry2Aa toxin.

A new chapter for anti-idiotypes in low molecular weight compound immunoassays

“…The successful isolation of Ab2s from scFv libraries can be improved. For example, a study using an Ab1 for the protein toxin Cry2Aa rapidly isolated eight Ab2s from scFvs libraries that they had previously only isolated two clones from, after using a chain shuffling technique [37]. However, this is an extra stage and complicates the process, which can clearly be simplified by using naïve nanobody libraries.…”

Anti-idiotypic VHH mediated environmentally friendly immunoassay for citrinin without mycotoxin

“…Thus, in comparison with immunoassays based on traditional chemically synthesized antigens, novel developed detection methods employing biosynthetic antigens of VHHs display preferable cost advantages and product stability, so they are suitable for commercial production and widespread use. Accordingly, several anti-idiotypic VHHs (AI-VHHs), including those for mycotoxin zearalenone, fumonisin B 1 , aflatoxin B 1 , Ochratoxin A, deoxynivalenol (DON), and other toxins, have been developed as toxin surrogates in immunoassays (Hao et al, 2020;Liu et al, 2018;Qiu et al, 2016;Shu et al, 2019;Wang et al, 2013;Wang et al, 2016;Zhang et al, 2019). However, these determination methods for mycotoxin surveillance only avoid the application one of the chemically synthetized mycotoxin conjugates or mycotoxin detection standard.…”

Development of a noncompetitive idiometric nanobodies phage immumoassay for the determination of fumonisin B 1

“…Ab2 is the second antibody that recognizes different epitopes within the hypervariable region of the primary antibody (Jerne, 1974). Alphatype Ab2 (Ab2α) and betatype Ab2 (Ab2β) are the two main types of anti-idiotypic antibodie (Lan et al, 2017;Liu et al, 2018). Ab2α recognizes an epitope, which is not part of the binding site (paratope) of the primary antibody, involved in antigen recognition.…”