The effect of varying the peptide linker length in a single chain variable fragment antibody against wogonin glucuronide

Paudel, Madan Kumar; Sakamoto, Seiichi; Huy, Le Van; Tanaka, Hiroyuki; Miyamoto, Tomofumi; Morimoto, Satoshi

doi:10.1016/j.jbiotec.2017.04.002

Cited by 7 publications

(5 citation statements)

References 26 publications

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“…The Fc region of immunoglobulin (MAb/PAb) is typically used as the epitope of secondary antibody for high versatility, while tags such as poly His-tag, T7-tag, and E-tag are commonly used as epitopes of secondary antibodies for rAb because they can be genetically incorporated into genes without disturbing the tertiary structure and activity of the rAb. Thus far, various scFvs against plant secondary metabolites have been constructed and expressed in E. coli to develop icELISA, including plumbagin [ 61 ], G-Re [ 62 ], DZ [ 63 ], wogonin glucuronide [ 64 ], and paclitaxel [ 65 ]. Similarly, Fab-based icELISA has been reported for artemisinin, which is produced from traditional Chinese herbal medicines, e.g., Artemisia annua L. and wogonin glucuronide, for their determination [ 66 , 67 ].…”

Section: Elisa For Plant Secondary Metabolitesmentioning

confidence: 99%

Enzyme-linked immunosorbent assay for the quantitative/qualitative analysis of plant secondary metabolites

et al. 2017

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Immunoassays are antibody-based analytical methods for quantitative/qualitative analysis. Since the principle of immunoassays is based on specific antigen–antibody reaction, the assays have been utilized worldwide for diagnosis, pharmacokinetic studies by drug monitoring, and the quality control of commercially available products. Berson and Yalow were the first to develop an immunoassay, known as radioimmunoassay (RIA), for detecting endogenous plasma insulin [1], a development for which Yalow was awarded the Nobel Prize in Physiology or Medicine in 1977. Even today, after half a century, immunoassays are widely utilized with some modifications from the originally proposed system, e.g., radioisotopes have been replaced with enzymes because of safety concerns regarding the use of radioactivity, which is referred to as enzyme immunoassay/enzyme-linked immunosorbent assay (ELISA). In addition, progress has been made in ELISA with the recent advances in recombinant DNA technology, leading to increase in the range of antibodies, probes, and even systems. This review article describes ELISA and its applications for the detection of plant secondary metabolites.

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Section: Elisa For Plant Secondary Metabolitesmentioning

confidence: 99%

Enzyme-linked immunosorbent assay for the quantitative/qualitative analysis of plant secondary metabolites

et al. 2017

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“…Varying the length of the linker peptide also affected scFv characterization. Shortening the linker of the VH-VL oriented scFv may be attractive in some important properties [16][17][18]. Our work obtained two scFvs against IL1RAP that were both unexpectedly mutated to a short sequence of linker peptide ( Figure 3A).…”

Section: Selection Of Yeast-displayed Candidate Anti-il1rap Scfvs Formentioning

confidence: 96%

“…Interestingly, both scFvs against IL1RAP contained a short sequence of Gly 4 Ser in linker peptide instead of a Gly 4 Ser repeat. Previous study indicated that the linker peptide between variable domains of heavy (VH) and light (VL) chains containing (Gly 4 Ser) n is one of the important factors that influence the binding activity and specificity of scFv against the target antigen [15,16]. Varying the length of the linker peptide also affected scFv characterization.…”

Section: Selection Of Yeast-displayed Candidate Anti-il1rap Scfvs Formentioning

confidence: 99%

Selection of novel human scFvs against cancer antigen IL1RAP by phage and yeast surface display technology

Duan

Jia

Xie

et al. 2020

Biotechnology & Biotechnological Equipment

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The objective of this study was isolation of humanized single-chain variable fragment (scFv) antibodies against IL1RAP, a promising cancer marker in leukemia and other tumors. Here, we screened for antibody fragments that recognized IL1RAP using a phage-display library of humanized scFv through several rounds of bio-panning and further FACS selection of enriched yeast-display sub-library. Two specific scFv clones 02 and 61 were isolated with high affinity against IL1RAP. Selected IL1RAP scFv antibodies containing Fc segment were expressed and purified from 293F cells. scFv-02 expressed and purified from mammalian cells exhibited higher IL1RAP binding affinity on IL1RAP-positive 293F cell surface compared to scFv-61. In addition, scFv-02 was significantly specific to IL1RAP, as no binding signal of IL1RAP was detected on the surface of IL1RAP CRISPR-Cas9 edited cell line generated in this study. Two scFv antibodies targeting IL1RAP were isolated by phage and yeast display screening. scFv-02 could serve as a promising material for further development of IL1RAP targeted immunotherapy of leukemia and other diseases. Our work also modified the process for the selection of scFv against targeted antigen by stepwise using phage display, yeast display and mammalian cell display techniques.

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“…Single‐chain variable fragment (scFv) antibody is an engineered antibody consisting of variable regions of heavy (VH) and light (VL) chains linked together with a flexible peptide linker (Figure ). Therefore, scFv‐based ELISA for determination of various plant bioactive compounds have been successfully developed . Application of scFv antibody in the development of immunoassay to determine plant bioactive compounds can be broadened by fusion with various protein tag such as green fluorescent protein (GFP) .…”

Section: Introductionmentioning

confidence: 99%

“…Therefore, scFv-based ELISA for determination of various plant bioactive compounds have been successfully developed. [22][23][24][25][26] Application of scFv antibody in the development of immunoassay to determine plant bioactive compounds can be broadened by fusion with various protein tag such as green fluorescent protein (GFP). 27,28 In this study, we constructed scFv antibody targeting miroestrol and successfully express the functional anti-miroestrol scFv antibody in the cytoplasm of Escherichia coli Shuffle T7.…”

mentioning

confidence: 99%

Preparation of a highly specific single chain variable fragment antibody targeting miroestrol and its application in quality control of Pueraria candollei by enzyme‐linked immunosorbent assay

Pongkitwitoon

Boonsnongcheep

Kitisripanya

et al. 2019

Phytochemical Analysis

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Introduction Miroestrol is the potent phytoestrogen isolated from White Kwao Krua (Pueraria candollei var. mirifica (Airy Shaw & Suvat.) Niyomdham, a Thai traditional medicinal plant. Nowadays, various health supplementary products featuring White Kwao Krua are available worldwide. A sensitive and rapid analytical method for quantification of miroestrol is necessary for quality control of these products. Objectives To prepare a single‐chain variable fragment (scFv) antibody specific to miroestrol and develop a scFv‐based enzyme‐linked immunosorbent assay (ELISA) for quantitative analysis of miroestrol in plant materials and health supplementary products. Methods A gene encoding anti‐miroestrol scFv antibody was constructed and expressed in Escherichia coli SHuffle T7 strain. Anti‐miroestrol scFv antibody was characterised and applied to ELISA. The developed scFv‐based ELISA method was validated for its sensitivity, specificity, accuracy and precision. Results Anti‐miroestrol scFv antibody was highly specific to miroestrol. The scFv‐based ELISA was applied to determine miroestrol in the range 0.06–7.81 μg/mL, with the limit of quantification of 0.06 μg/mL miroestrol. The accuracy of the assay was validated by its 95.08–103.99% recovery from the spiked miroestrol recovery experiment and in good correlation with the results from the monoclonal antibody‐based ELISA. The relative standard deviation of the intra‐ and inter‐assay were less than 6.0%. Conclusion The developed scFv‐based ELISA was sensitive, specific, accurate, and precise for determination of miroestrol and useful for quality control of P. candollei plant raw materials and supplementary products.

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The effect of varying the peptide linker length in a single chain variable fragment antibody against wogonin glucuronide

Cited by 7 publications

References 26 publications

Enzyme-linked immunosorbent assay for the quantitative/qualitative analysis of plant secondary metabolites

Enzyme-linked immunosorbent assay for the quantitative/qualitative analysis of plant secondary metabolites

Selection of novel human scFvs against cancer antigen IL1RAP by phage and yeast surface display technology

Preparation of a highly specific single chain variable fragment antibody targeting miroestrol and its application in quality control of Pueraria candollei by enzyme‐linked immunosorbent assay

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