Increased Sensitivity of Cholera Toxin Detection by Enzyme‐Linked Immunosorbent Assay with Chemical Immobilization of Antibody onto Nylon Surface Providing Hydrophobicity

Koike, Norio; Yabushita, Yasunori; Yan, Zhang Dong; Iida, Tetsuya; Honda, Takeshi

doi:10.1111/j.1348-0421.1999.tb01235.x

Cited by 1 publication

(1 citation statement)

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“…For example, scFvs against SEB showed no cross-reactivity versus other antigens in the panel, and BIAcore analysis indicated a K d ϭ 7.8 nM for scFv SEB13, which was comparable to anti-SEB murine mAbs obtained from a secondary immune response requiring intrasplenic boosting (55). Also, a high-affinity scFv against CTB was obtained, with the K d value the first reported to date, and the scFv affinity was estimated to exceed that of previously isolated murine mAbs from immunization (56,57). The exquisite specificity of the scFvs between the highly homologous ricin agglutinins, RCA 60 and RCA 120 , was also notable, and such scFvs should be useful for the detection and distinction of these toxins.…”

Section: Discussionmentioning

confidence: 74%

A method for the generation of combinatorial antibody libraries using pIX phage display

Gao

Mao

Kaufmann

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

117

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For more than a decade, phage displayed combinatorial antibody libraries have been used to generate and select a wide variety of antibodies. We previously reported that the phage coat proteins pVII and pIX could be used to display the heterodimeric structure of the antibody Fv region. Herein, aspects of this technology were invoked and extended to construct a large, human single-chain Fv (scFv) library of 4.5 ؋ 10 9 members displayed on pIX of filamentous bacteriophage. Furthermore, the diversity, quality, and utility of the library were demonstrated by the selection of scFv clones against six different protein antigens. Notably, more than 90% of the selected clones showed positive binding for their respective antigens after as few as three rounds of panning. Analyzed scFvs were also found to be of high affinity. For example, kinetic analysis (BIAcore) revealed that scFvs against staphylococcal enterotoxin B and cholera toxin B subunit had a nanomolar and subnanomolar dissociation constant, respectively, affording affinities comparable to, or exceeding that, of mAbs obtained from immunization. High specificity was also attained, not only between very distinct proteins, but also in the case of the Ricinus communis (''ricin'') agglutinins (RCA60 and RCA120), despite >80% sequence homology between the two. The results suggested that the performance of pIX-display libraries can potentially exceed that of the pIII-display format and make it ideally suited for panning a wide variety of target antigens.

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Section: Discussionmentioning

confidence: 74%