Citation: CAO Dong-Mei, XU Yang, TU Zhui, LI Yan-Ping, XIONG Liang, FU Jin-Heng. One-step Enzyme Linked Immunosorbent Assay for Detection of Aflatoxin B₁ Using a Nanobody-Alkaline Phosphatase Fusion Protein[J]. Chinese Journal of Analytical Chemistry, ;2016, 44(7): 1085-1091. doi: 10.11895/j.issn.0253-3820.150902 shu

One-step Enzyme Linked Immunosorbent Assay for Detection of Aflatoxin B₁ Using a Nanobody-Alkaline Phosphatase Fusion Protein

CAO Dong-Mei ^1, ,
XU Yang ^1,2 ,
TU Zhui ¹ ,
LI Yan-Ping ² ,
XIONG Liang ^1,3 ,
FU Jin-Heng ²

1.
State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang 330047, China;
2.
Jiangxi-OAI Joint Research Institute, Nanchang University, Nanchang 330047, China;
3.
Department of Preventive Medicine, Gannan Medical College, Ganzhou 341000, China

Received Date: 13 November 2015
Revised Date: 8 April 2016

Fund Project: This work was supported by the National Natural Science Foundation of China (No. 31301479), the Research Foundation for Young Scientists of State Key Laboratory of Food Science and Technology (No. SKLF-QN-201513), and the Major Program of Natural Science Foundation of Jiangxi Province (20152ACB20005)

Figures(5)

A phage clone (designated as the anti-aflatoxin B₁ nanobody, G8) specific binding to aflatoxin B₁ (AFB₁) was screened from an immunized nanobody phage library. The DNA fragment encoding G8 was subcloned into the vector pET25b(+)-alkaline phosphatase (AP), and fused to the N terminal of AP. The fusion protein G8-AP was expressed in E. coli BL21 (Rosetta, DE3) as soluble protein, and purified by immobilized metal affinity chromatography. The purified G8-AP exhibited AP activity of 364.5±8.3 U/mg. Enzyme-linked immunosorbent assay (ELISA) showed that G8-AP was also capable of recognizing AFB₁, with negligible cross-reactivity toward other six common mycotoxins. A one-step ELISA was established to detect AFB₁ using G8-AP. The effects of methanol concentration, ionic strength, and pH on the detection sensitivity of the G8-AP-ELISA were evaluated and optimized. Under the optimized conditions (methanol 20%, NaCl 20 mmol/L, pH 7.4), the half inhibitory concentration (IC₅₀) of the G8-AP based ELISA was 19.8 ng/mL, with the linear range of 4.3-92 ng/mL and the detection limit of 2.6 ng/mL. The recoveries of AFB₁ for corn and wheat samples were 90.4%-101.5%.
- Anti-aflatoxin B₁ Nanobody,
- Alkaline phosphatase,
- Fusion expression,
- Enzyme-linked immunosorbent assay
1. [1]
2. [2]
3. [3]
4. [4]
5. [5]
6. [6]
7. [7]
8. [8]
9. [9]
10. [10]
11. [11]
12. [12]
13. [13]
14. [14]
15. [15]
16. [16]
17. [17]
18. [18]
19. [19]
20. [20]
21. [21]
22. [22]
23. [23]
24. [24]
25. [25]
26. [26]
27. [27]
28. [28]
29. [29]
1. [1]
  Yuyang Xu , Ruying Yang , Yanzhe Zhang , Yandong Liu , Keyi Li , Zehui Wei . Research Progress of Aflatoxins Removal by Modern Optical Methods. University Chemistry, 2024, 39(11): 174-181. doi: 10.12461/PKU.DXHX202402064
2. [2]
  Zhaoyang Li , Haiyan Zhao , Yali Zhang , Yuan Zhang , Shiqiang Cui . Integration of Nobel Prize Achievements in Analytical Technology with College Instrumental Analysis Course. University Chemistry, 2025, 40(3): 269-276. doi: 10.12461/PKU.DXHX202405131
3. [3]
  Jingyi Chen , Fu Liu , Tiejun Zhu , Kui Cheng . Practice of Integrating Ideological and Political Education into Raman Spectroscopy Analysis Experiment Course. University Chemistry, 2024, 39(2): 140-146. doi: 10.3866/PKU.DXHX202310111
4. [4]
  Congying Wen , Zhengkun Du , Yukun Lu , Zongting Wang , Hua He , Limin Yang , Jingbin Zeng . Teaching Reform and Practice of Modern Analytical Technology under the Integration of Science, Industry, and Education. University Chemistry, 2024, 39(8): 104-111. doi: 10.3866/PKU.DXHX202312089
5. [5]
  Tianlong Zhang , Rongling Zhang , Hongsheng Tang , Yan Li , Hua Li . Exploration on the Integration Mode of Instrumental Analysis with Science and Education under the Background of Artificial Intelligence Era. University Chemistry, 2024, 39(8): 365-374. doi: 10.12461/PKU.DXHX202403014
6. [6]
  Linlin Guo , Jinjun Zhang , Chengpeng Miao , Bojing Liu , Xiaozhen Fan . Design and Practice of Integrating Ideological and Political Education into Instrumental Analysis Course Based on OBE Concept: Introduction. University Chemistry, 2024, 39(11): 87-95. doi: 10.12461/PKU.DXHX202403001
7. [7]
  Liwei Wang , Guangran Ma , Li Wang , Fugang Xu . A Comprehensive Analytical Chemistry Experiment: Colorimetric Detection of Vitamin C Using Nanozyme and Smartphone. University Chemistry, 2024, 39(8): 255-262. doi: 10.3866/PKU.DXHX202312094
8. [8]
  Kuaibing Wang , Honglin Zhang , Wenjie Lu , Weihua Zhang . Experimental Design and Practice for Recycling and Nickel Content Detection from Waste Nickel-Metal Hydride Batteries. University Chemistry, 2024, 39(11): 335-341. doi: 10.12461/PKU.DXHX202403084
9. [9]
  Miao-Miao Chen , Min-Ling Zhang , Xiao Song , Jun Jiang , Xiaoqian Tang , Qi Zhang , Xiuhua Zhang , Peiwu Li . Smartphone-assisted electrochemiluminescence imaging test strips towards dual-signal visualized and sensitive monitoring of aflatoxin B1 in corn samples. Chinese Chemical Letters, 2025, 36(1): 109785-. doi: 10.1016/j.cclet.2024.109785
10. [10]
  Haiyuan Wang , Yiming Tang , Haoran Guo , Guohui Chen , Yajing Sun , Chao Zhao , Zhen Zhang . Comprehensive Chemistry Experimental Teaching Design Based on the Integration of Science and Education: Preparation and Catalytic Properties of Silver Nanomaterials. University Chemistry, 2024, 39(10): 219-228. doi: 10.12461/PKU.DXHX202404067
11. [11]
  Jie ZHAO , Sen LIU , Qikang YIN , Xiaoqing LU , Zhaojie WANG . Theoretical calculation of selective adsorption and separation of CO₂ by alkali metal modified naphthalene/naphthalenediyne. Chinese Journal of Inorganic Chemistry, 2024, 40(3): 515-522. doi: 10.11862/CJIC.20230385
12. [12]
  Simin Fang , Wei Huang , Guanghua Yu , Cong Wei , Mingli Gao , Guangshui Li , Hongjun Tian , Wan Li . Integrating Science and Education in a Comprehensive Chemistry Design Experiment: The Preparation of Copper(I) Oxide Nanoparticles and Its Application in Dye Water Remediation. University Chemistry, 2024, 39(8): 282-289. doi: 10.3866/PKU.DXHX202401023
13. [13]
  Junli Liu . Practice and Exploration of Research-Oriented Classroom Teaching in the Integration of Science and Education: a Case Study on the Synthesis of Sub-Nanometer Metal Oxide Materials and Their Application in Battery Energy Storage. University Chemistry, 2024, 39(10): 249-254. doi: 10.12461/PKU.DXHX202404023
14. [14]
  Chunmei GUO , Weihan YIN , Jingyi SHI , Jianhang ZHAO , Ying CHEN , Quli FAN . Facile construction and peroxidase-like activity of single-atom platinum nanozyme. Chinese Journal of Inorganic Chemistry, 2024, 40(9): 1633-1639. doi: 10.11862/CJIC.20240162
15. [15]
  Fang Niu , Rong Li , Qiaolan Zhang . Analysis of Gas-Solid Adsorption Behavior in Resistive Gas Sensing Process. University Chemistry, 2024, 39(8): 142-148. doi: 10.3866/PKU.DXHX202311102
16. [16]
  Zhuoya WANG , Le HE , Zhiquan LIN , Yingxi WANG , Ling LI . Multifunctional nanozyme Prussian blue modified copper peroxide: Synthesis and photothermal enhanced catalytic therapy of self-provided hydrogen peroxide. Chinese Journal of Inorganic Chemistry, 2024, 40(12): 2445-2454. doi: 10.11862/CJIC.20240194
17. [17]
  Jingke LIU , Jia CHEN , Yingchao HAN . Nano hydroxyapatite stable suspension system: Preparation and cobalt adsorption performance. Chinese Journal of Inorganic Chemistry, 2024, 40(9): 1763-1774. doi: 10.11862/CJIC.20240060
18. [18]
  Peiqi Gao , Jiao Zheng , LiMiao Chen , Yi Zhang . Exploration of the Deep Integration Strategy between Innovation and Entrepreneurship Education and Applied Chemistry Major Courses. University Chemistry, 2024, 39(6): 214-219. doi: 10.3866/PKU.DXHX202310086
19. [19]
  Yecang Tang , Shan Ling , Zhen Fang . Exploration of a Hierarchical and Integration-Oriented Talent Training Model in the Demonstration Center for Experimental Chemistry Education. University Chemistry, 2024, 39(7): 188-192. doi: 10.12461/PKU.DXHX202405107
20. [20]
  Hengwei Wei , Liqiu Zhao , Jiqiang Geng , Xuebo Xu , Yingpeng Ma , Yuhao Liu , Mingzhe Han , Huan Jiao , Lingling Wei . Research on Safety Management of Hazardous Chemicals and Talent Cultivation in Universities Driven by Production-Education Integration. University Chemistry, 2024, 39(10): 289-298. doi: 10.12461/PKU.DXHX202403022

Get Citation

PDF

XML

Metrics

PDF Downloads(16)
Abstract views(1519)
HTML views(242)

通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

One-step Enzyme Linked Immunosorbent Assay for Detection of Aflatoxin B1 Using a Nanobody-Alkaline Phosphatase Fusion Protein

Metrics

通讯作者: 陈斌, bchen63@163.com

One-step Enzyme Linked Immunosorbent Assay for Detection of Aflatoxin B₁ Using a Nanobody-Alkaline Phosphatase Fusion Protein