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Development of an Indirect Competitive ELISA Kit for the Detection of Soybean Allergenic Protein Gly m Bd 28K

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Abstract

The complementary DNA (cDNA) sequence of Gly m Bd 28K was expressed in Escherichia coli BL21 (DE3) as an inclusion body under the induction of 1.0 mmol/L of isopropyl β-D-thiogalactoside (IPTG). The inclusion bodies were solubilized in 6 M guanidine hydrochloride and refolded by rapid dilution. After renaturation, the purity of the recovered recombinant protein was up to 90 %, and its molecular weight was 29.1 kDa. A monoclonal antibody that was specific for recombinant Gly m Bd 28K protein was produced and characterized. Based on the mAb, an indirect competitive enzyme-linked immunosorbent kit (icELISA kit) for the specific detection of recombinant Gly m Bd 28K protein was developed, which showed that the IC50 was 3.19 μg/L and the sensitivity was 0.235 μg/L. The average relative standard deviation (RSD) was 10.28 %, and the average recovery was 96.73 %, which indicated that the kit was highly sensitive and accurate. The icELISA kit remained usable for more than 6 months when stored at 4 °C. Therefore, the icELISA is a valuable tool for the rapid and sensitive detection of Gly m Bd 28K in food and feed products from soybeans.

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Acknowledgments

This study was financially supported by the Natural Science Foundation of China (NSFC, 31301409), Plan of Nature Science Fundamental Research in Henan University of Technology (2014JCYJ01), and Key Scientific Research Projects of Henan colleges and Universities (16A550001).

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Authors and Affiliations

  1. School of Food Science and Technology, Henan University of Technology, Zhengzhou, 450001, Henan, China

    Jun Xi & Qiaoqiao Shi

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  1. Jun Xi
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  2. Qiaoqiao Shi
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Correspondence to Jun Xi.

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Jun Xi declares she has no conflict of interest. Qiaoqiao Shi declares she has no conflict of interest. This article does not contain any studies with human subjects.

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The studies with animals were performed following all institutional and national guidelines for the care and use of laboratory animals.

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Xi, J., Shi, Q. Development of an Indirect Competitive ELISA Kit for the Detection of Soybean Allergenic Protein Gly m Bd 28K. Food Anal. Methods 9, 2998–3005 (2016). https://doi.org/10.1007/s12161-016-0493-7

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  • DOI: https://doi.org/10.1007/s12161-016-0493-7

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