Enzyme‐Linked Immunosorbent Assay for the Determination of Total Ginsenosides in Ginseng

Morinaga, Osamu; Tanaka, Hiroyuki; Shoyama, Yukihiro

doi:10.1080/00032710500476979

Cited by 13 publications

(12 citation statements)

References 17 publications

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“…5). It is noted that the assay using GRe-scFv displayed almost the same sensitivity as that using MAb-4G10 [8].…”

Section: Methods Validationmentioning

confidence: 84%

“…To investigate the accuracy of this assay, the total ginsenoside concentrations of various ginseng samples determined by ELISA using GRe-scFv were compared with those determined by ELISA using MAb-4G10, which showed a high correlation with HPLC analysis in our previous report [8]. Table 3 shows the results of quantitative ELISA for total ginsenosides using GRe-scFv and MAb-4G10.…”

Section: Methods Validationmentioning

confidence: 99%

“…In this assay, G-Re-human serum albumin conjugates (GRe-HSA), which were revealed to have 5 molecules of G-Re per 1 molecule of HSA in our previous study [8], were used as the solidphase antigen. A 96-well immunoplate (Nunc, Roskilde, Denmark) was coated with 100 ll of GRe-HSA (2 lg/ml) in 50 mM carbonate buffer (pH 9.6) and incubated for 1 h. After washing the plate, it was treated with 300 ll of phosphate-buffered saline containing 10% (w/v) skimmed milk for 1 h to reduce nonspecific adsorption.…”

Section: Refolding Of Gre-scfv Proteinmentioning

confidence: 99%

“…The MAb against G-Re (MAb-4G10) was shown to be an effective tool for ELISA for determining total ginsenosides in ginseng because of its wide cross-reactivities (CR) with 20(S)-protopanaxadiol-and 20(S)-protopanaxatriol-type ginsenosides [8], and the assay showed potential as a fast and reliable method for assessing the total ginsenosides concentration of plant samples. However, it is time-consuming and labor-intensive to obtain MAb.…”

Section: Introductionmentioning

confidence: 99%

“…In our previous studies, we prepared monoclonal antibodies (MAb) against ginsenoside Re (G-Re) [5], ginsenoside Rb1 (G-Rb1) [6], and ginsenoside Rg1 (G-Rg1) [7] for development of enzyme-linked immunosorbent assays (ELISA) [8,9] and chromatographic immunostaining methods [5,10] for the determination of various ginsenosides. The MAb against G-Re (MAb-4G10) was shown to be an effective tool for ELISA for determining total ginsenosides in ginseng because of its wide cross-reactivities (CR) with 20(S)-protopanaxadiol-and 20(S)-protopanaxatriol-type ginsenosides [8], and the assay showed potential as a fast and reliable method for assessing the total ginsenosides concentration of plant samples.…”

Section: Introductionmentioning

confidence: 99%

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Single-chain variable fragment antibody against ginsenoside Re as an effective tool for the determination of ginsenosides in various ginsengs

et al. 2010

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A single-chain variable fragment antibody (scFv) against ginsenoside Re (G-Re) was constructed and applied to an enzyme-linked immunosorbent assay (ELISA) for determining the total concentration of ginsenosides in various ginsengs. The variable heavy and light chain genes were cloned directly from the cDNA of the 4G10 hybridoma cell line and assembled by means of splicing by overlapping extension PCR (SOE-PCR) using specific primers designed to have flexible peptide (Gly(4)Ser)(3) between the variable heavy chain and light chain domains. The constructed scFv gene was ligated into the pET28a expression vector and transformed into E. coli BL21 (DE3). The recombinant scFv against G-Re (GRe-scFv) was expressed as a chimera protein containing the His6-tag at its N-termini, purified by immobilized metal ion affinity chromatography (IMAC), and refolded by a stepwise dialysis method. The yield of GRe-scFv after purification was 1.7 mg per liter of culture medium. Characterization of GRe-scFv revealed that it retained the characteristics of the parental monoclonal antibody (MAb) against G-Re (MAb-4G10) which has wide cross-reactivity with 20(S)-protopanaxadiol- and 20(S)-protopanaxatriol-type ginsenosides. The detectable range for G-Re in ELISA using scFv antibody was 0.02-10 µg/ml. Based on validation analysis, the use of GRe-scFv in ELISA is a precise, accurate, and sensitive method. In light of the time-consuming and labor-intensive procedures for the preparation of MAb, speedy bacterial expression of GRe-scFv is a powerful alternative tool for producing MAb to use in ELISA for quantitative analysis of total ginsenoside concentrations.

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“…5). It is noted that the assay using GRe-scFv displayed almost the same sensitivity as that using MAb-4G10 [8].…”

Section: Methods Validationmentioning

confidence: 84%

Section: Methods Validationmentioning

confidence: 99%

Section: Refolding Of Gre-scfv Proteinmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Single-chain variable fragment antibody against ginsenoside Re as an effective tool for the determination of ginsenosides in various ginsengs

et al. 2010

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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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Lateral flow immunoassay for small-molecules detection in phytoproducts: a review

et al. 2022

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Phytoproducts are involved in various fields of industry. Small-molecule (Mw < 900 Da) organic compounds can be used to indicate the quality of plant samples in the perspective of efficacy by measuring the necessary secondary metabolites and in the perspective of safety by measuring the adulterant level of toxic compounds. The development of reliable detection methods for these compounds in such a complicated matrix is challenging. The lateral flow immunoassay (LFA) is one of the immunoassays well-known for its simplicity, portability, and rapidity. In this review, the general principle, components, format, and application of the LFA for phytoproducts are discussed.

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Enzyme‐Linked Immunosorbent Assay for the Determination of Total Ginsenosides in Ginseng

Cited by 13 publications

References 17 publications

Single-chain variable fragment antibody against ginsenoside Re as an effective tool for the determination of ginsenosides in various ginsengs

Single-chain variable fragment antibody against ginsenoside Re as an effective tool for the determination of ginsenosides in various ginsengs

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

Lateral flow immunoassay for small-molecules detection in phytoproducts: a review

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