Human 4F5 single-chain Fv antibody recognizing a conserved HA1 epitope has broad neutralizing potency against H5N1 influenza A viruses of different clades

Zhang, Xiao; Qi, Xian; Zhang, Qianqian; Zeng, Xianying; Shi, Zhiyang; Qiu, Jin; Zhan, Feng; Xu, Yan; Liu, Zhe; Feng, Zijian; Jiao, Yongjun

doi:10.1016/j.antiviral.2013.05.001

Cited by 17 publications

(24 citation statements)

References 45 publications

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“…Previously, two mAbs H5M9 [14] and 4F5 [15], were reported to interact with amino acid residues close to 9F4's binding site. H5M9 was produced by immunising mice with HA protein of A/Goose/Guangdong/1/96 (H5N1).…”

Section: Generation Of Neutralising Mabs Binding To the Ve Subdomainmentioning

confidence: 99%

“…Interestingly, several mAbs in this class have been documented to bind to the vestigial esterase (VE) subdomain of HA of the H5N1 subtype and they have been shown to bind to multiple clades of H5N1 viruses [11][12][13][14][15]. However, they typically do not bind to other subtypes of IAV which suggests that the VE subdomain is conserved within a subtype but diverse among subtypes.…”

Section: Introductionmentioning

confidence: 99%

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The Vestigial Esterase Domain of Haemagglutinin of H5N1 Avian Influenza A Virus: Antigenicity and Contribution to Viral Pathogenesis

Zheng

Paul

et al. 2018

Preprint

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Initial attempts to develop monoclonal antibodies as therapeutics to resolve influenza infections focused mainly on searching for antibodies with the potential to neutralise the virus in vitro with classical haemagglutination inhibition and micro-neutralisation assays. This led to the identification of many antibodies that bind to the head domain of haemagglutinin (HA) which generally have potent neutralisation capabilities that block viral entry or viral membrane fusion. However, this class of antibodies has a narrow breadth of protection in that they are usually strain specific. This led to the emphasis on stalk targeting antibodies which are able to bind a broad range of viral targets that span across different influenza subtypes.Preprints (www.preprints.org) | NOT PEER-REVIEWED | Posted:

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Section: Generation Of Neutralising Mabs Binding To the Ve Subdomainmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

The Vestigial Esterase Domain of Haemagglutinin of H5N1 Avian Influenza A Virus: Antigenicity and Contribution to Viral Pathogenesis

Zheng

Paul

et al. 2018

Preprint

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“…However, such result does not exclude that scFv recognized the same epitope as mAb. To prevent hemagglutination, antibody must bind to or near RBS and cause steric hindrance (Ascione et al, 2009;Zhang et al, 2013). It is more likely that the reduced size of scFv is not an effective barrier between virus and the red cell.…”

Section: Construction and Expression Of The Scfv -Mab6-9-1 Derivativementioning

confidence: 99%

Characterization of mAb6-9-1 monoclonal antibody against hemagglutinin of avian influenza virus H5N1 and its engineered derivative, single-chain variable fragment antibody

Sawicka¹,

Siedlecki²,

Kalenik³

et al. 2016

Acta Biochim Pol

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Hemagglutinin (HA), as a major surface antigen of influenza virus, is widely used as a target for production of neutralizing antibodies. Monoclonal antibody, mAb6-9-1, directed against HA of highly pathogenic avian influenza virus A/swan/Poland/305-135V08/2006(H5N1) was purified from mouse hybridoma cells culture and characterized. The antigenic specificity of mAb6-9-1 was verified by testing its cross-reactivity with several variants of HA. The mimotopes recognized by mAb6-9-1 were selected from two types of phage display peptide libraries. The comparative structural model of the HA variant used for antibody generation was developed to further facilitate epitope mapping. Based on the sequences of the affinity-selected polypeptides and the structural model of HA the epitope was located to the region near the receptor binding site (RBS). Such localization of the epitope recognized by mAb6-9-1 is in concordance with its moderate hemagglutination inhibiting activity and its antigenic specificity. Additionally, total RNA isolated from the hybridoma cell line secreting mAb6-9-1 was used for obtaining two variants of cDNA encoding recombinant single-chain variable fragment (scFv) antibody. To ensure high production level and solubility in bacterial expression system, the scFv fragments were produced as chimeric proteins in fusion with thioredoxin or displayed on a phage surface after cloning into the phagemid vector. Specificity and affinity of the recombinant soluble and phage-bound scFv were assayed by suitable variants of ELISA test. The observed differences in specificity were discussed.

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“…Each well was blocked with blocking buffer (Table 3) at 4°C overnight. Then incubated scFv antibodies doubling diluted from 1:10 to 1:20480 in blocking buffer at 37°C for 2 h. ScFv 4F5 (Zhang et al, 2013) produced in our lab was used as negative control. Unbound antibodies were removed by washing 3 times with 0.05% PBST.…”

Section: Elisa Assaymentioning

confidence: 99%

“…The screening procedures and bacteria culture as well as transformation were operated as described elsewhere (Zhang et al, 2013). After four rounds of selection, E. coli XL1-Blue infected with phage were plated on agar plates and incubated overnight at 37°C.…”

Section: Panning and Expression Of Anti-znt8 Scfvmentioning

confidence: 99%

Screening and identification of human ZnT8-specific single-chain variable fragment (scFv) from type 1 diabetes phage display library

Wang

et al. 2016

Sci. China Life Sci.

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Zinc transporter 8 (ZnT8) is a major autoantigen and a predictive marker in type 1 diabetes (T1D). To investigate ZnT8-specific antibodies, a phage display library from T1D was constructed and single-chain antibodies against ZnT8 were screened and identified. Human T1D single-chain variable fragment (scFv) phage display library consists of approximately 110 8 clones. After four rounds of bio-panning, seven unique clones were positive by phage ELISA. Among them, C27 and C22, which demonstrated the highest affinity to ZnT8, were expressed in Escherichia coli Top10F' and then purified by affinity chromatography. C27 and C22 specifically bound ZnT8 N/C fusion protein and ZnT8 C terminal dimer with one Arg325Trp mutation. The specificity to human islet cells of these scFvs were further confirmed by immunohistochemistry. In conclusion, we have successfully constructed a T1D phage display antibody library and identified two ZnT8-specific scFv clones, C27 and C22. These ZnT8-specific scFvs are potential agents in immunodiagnostic and immunotherapy of T1D.Zinc transporter 8 (ZnT8), phage display, single-chain variable fragment (scFv), type 1 diabetes (T1D) Citation:Wu, Q., Wang, X., Gu, Y., Zhang, X., Qin, Y., Chen, H., Xu, X., Yang, T., and Zhang, M. (2016). Screening and identification of human ZnT8-specific single-chain variable fragment (scFv) from type 1 diabetes phage display library. Sci China Life Sci 59, 686 -693.

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Human 4F5 single-chain Fv antibody recognizing a conserved HA1 epitope has broad neutralizing potency against H5N1 influenza A viruses of different clades

Cited by 17 publications

References 45 publications

The Vestigial Esterase Domain of Haemagglutinin of H5N1 Avian Influenza A Virus: Antigenicity and Contribution to Viral Pathogenesis

The Vestigial Esterase Domain of Haemagglutinin of H5N1 Avian Influenza A Virus: Antigenicity and Contribution to Viral Pathogenesis

Characterization of mAb6-9-1 monoclonal antibody against hemagglutinin of avian influenza virus H5N1 and its engineered derivative, single-chain variable fragment antibody

Screening and identification of human ZnT8-specific single-chain variable fragment (scFv) from type 1 diabetes phage display library

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