查找使用阿拉丁产品的科研文献

An ic-ELISA and immunochromatographic strip assay for the detection of 2,4-dichlorophenoxyacetic acid in bean sprouts and cabbage

Xiaomin Pan,Xinxin Xu,Shanshan Song,Liguang Xu,Hua Kuang,Xiaoling Wu,Liqiang Liu,Chuanlai Xu

The compound 2,4-dichlorophenoxyacetic acid (2,4-D) is a synthetic plant growth regulator , which is widely used in agricultural production. Consequently, it is necessary to establish a rapid and sensitive detection method to monitor its use and prevent the environmental and human health problems caused by overuse. In this study, a monoclonal antibody (mAb) 2D5 against 2,4-D was prepared, and based on it, an ic-ELISA and an immunochromatographic strip assay (ICA) were developed for rapid detection. The 50% inhibitory concentration (IC 50 ) of the mAb against 2,4-D was 1.7 ng mL −1 and the mAb showed no cross-reactivity with other plant growth regulators with similar structures to 2,4-D. Under optimum conditions, the ICA showed a visual detection limit (vLOD) of 20 ng mL −1 and a cut-off value of 200 ng mL −1 in bean sprouts, and cabbage samples gave the same results. The quantitative detection showed the linear detection ranges for bean sprouts and cabbage samples were 6.3–73.0 and 5.4–132.6 ng mL −1 , respectively. The ICA was successfully applied to detect 2,4-D in bean sprouts and cabbage samples and achieved good recoveries ranging from 94.7% to 104.3% with a coefficient of variation (CV) less than 8.3%. Hence, the ICA provides a sensitive, efficient and reliable detection method, which has broad application prospects in the detection of a large number of samples.

Development of ic-ELISA and lateral-flow immunochromatographic strip for detection of vitamin B2 in an energy drink and vitamin tablets

Lu Zeng,Wei Jiang,Liqiang Liu,Shanshan Song,Hua Kuang

Vitamin B2 (riboflavin) is a water-soluble vitamin that has important roles in human health. In this study, we developed a sensitive and specific monoclonal antibody-based indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and lateral-flow immunochromatographic assay (ICA) strip for the rapid detection of vitamin B2. Following routine fusion and selection, the optimum monoclonal antibody against vitamin B2 was obtained. The 50% inhibitory concentration and limit of detection of ic-ELISA were 8.18 and 1.80 ng/mL, respectively. The cut-off value of the lateral-flow ICA strip was 50 ng/mL. The results revealed that our developed methods are suitable for the on-site detection and mass screening of vitamin B2 in food and pharmaceutical products.

Development and characterisation of an ultrasensitive monoclonal antibody for chloramphenicol

Naifeng Xu,Liguang Xu,Wei Ma,Hua Kuang,Chuanlai Xu

In this study, a highly sensitive monoclonal antibody (McAb) against chloramphenicol (CAP) was successfully raised and characterised by indirect competitive enzyme-linked immunosorbent assay (ic-ELISA). Three antigens coupled with different proteins were synthesised and determined by the UV-vis method. After five rounds of immunisation, antisera were collected from six Balb/C mice and screened by ic-ELISA. The standard inhibition curve of one CAP McAb (labelled 6F11) showed the best sensitivity to CAP after optimisation of the principal working conditions. The half-maximal inhibition concentration (IC50) was calculated to be 0.01 ng/mL and the linear working range was from 0.0028 ng/mL to 0.040 ng/mL with a correlation coefficient of 0.996. This McAb showed high specificity, with negligible cross-reactivities detected towards CAP analogues (florfinicol, florfinicol amine, thiamphenicol). The recovery rate was 98.9–106.8% while the coefficient of variation was 4.45–6.05%. The IC50 value and other parameters obtained for this CAP McAb suggest that it is likely to have a promising future in further applications in combination with different analytical techniques.

Development of a highly sensitive immunoassay for detecting aminopyrine abuse in herbal tea

Zian Wang,Zhiwei Liu,Tian Guan,Xi Zeng,Runlin Shen,Zhaodong Li,Yi Lei,Zhenlin Xu,Zhili Xiao,Hongtao Lei,Dingqiang Huang

With the popularity of herbal tea in China, many food fraudsters have added illegal drugs to herbal tea to enhance its functions, among which aminopyrine is widely abused as an antipyretic and analgesic. Presently, there is no immunoassays for aminopyrine, and it is difficult to achieve real-time detection in the field. Based on a polyclonal antibody of aminopyrine with high specificity and sensitivity, an optimal combination of coating antigen/antibody was obtained by screening different coating antigens. On this basis, a sensitive ic-ELISA method was established to detect aminopyrine in herbal tea. The detection limit of the ic-ELISA was 0.18 ng mL−1, which was much lower than the 100 ng mL−1 required as a standard. The method had good consistency with LC-MS in the detection of actual samples and could be used as a reliable method for the detection of aminopyrine in herbal tea.

Development of an ic-ELISA and a TRFICA for the detection of fipronil with a new hapten design

Liu Junwei,Wu Yuqing,Liu Chenchen,Cai Yan,Zhang Rui,Lu Yang

Based on the new hapten of fipronil, an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and time-resolved fluorescent immunochromatographic assay (TRFICA) for the determination of fipronil were developed. The sensitivity (IC 50 ) of the ic-ELISA method was 2.33 ± 0.14 ng mL −1 (n = 3), and the limit of detection (LOD) (IC 20 ) was 0.27 ± 0.02 ng mL −1 (n = 3). The cross-reaction rates of the three fipronil metabolites were all higher than 66.7%, and those of the three analogs were no more than 0.05%. The TRFICA was optimized with a handheld reader. The LOD was 0.862 ng mL −1 (n = 3). In this study, the pretreatment methods of samples such as Chinese cabbage, spinach, apple and egg, were explored. Methanol and acetonitrile were used to extract different food samples and the diluents were optimized. Three concentration levels in the spiked food samples were detected by the two methods. The range of recovery was 88.65–116.32% for ic-ELISA and 95.48–112.61% for TRFICA. In this study, the accuracy of the two methods was also judged by testing commercial quality control samples. The results showed that the accuracy was 83.75%–113.33% for ic-ELISA and 98.33–107.13% for TRFICA.

Development of an ic-ELISA and immunochromatographic assay strip for the rapid detection of chloridazon in oranges and celery

Qing Zhang,Aihong Wu,Jinyan Li,Liqiang Liu,Hua Kuang,Chuanlai Xu,Lingling Guo

Chloridazon (CLZ) is a selective herbicide used in the control of annual broadleaf weeds. The misuse or abuse of CLZ may result in the accumulation of CLZ in crops and water, which can pose a risk to human health. In this study, a hapten of CLZ with three carbon spacer arms was designed and a highly sensitive and specific antibody against CLZ was prepared with a half-maximal inhibitory concentration of 0.630 ng mL−1 and a linear range of 0.181–2.195 ng mL−1.Based on this antibody, we developed an immunochromatographic assay (ICA) strip for the detection of CLZ in oranges and celery. Under optimized conditions, the visual limit of detection was 2 ng mL−1 and 10 ng mL−1 in oranges and celery, respectively, and the cut-off value was 50 ng mL−1. In CLZ-spiked samples and the recovery test, the results of the ICA strip were consistent with those of indirect competitive enzyme-linked immunosorbent assay (ic-ELISA). Therefore, the ICA strip developed in our study represents an efficient and reliable method for the rapid screening of CLZ in oranges and celery.

Development of an immunochromatographic test strip and ic-ELISA for tetrabromobisphenol: a detection in lake water and rice pudding samples

Li Yu,Liqiang Liu,Shanshan Song,Hua Kuang,Chuanlai Xu

Preparation of an Immunoaffinity Column Based on Bispecific Monoclonal Antibody for Aflatoxin B1 and Ochratoxin A Detection Combined with ic-ELISA

Disha Lu,Xu Wang,Ruijue Su,Yongjian Cheng,Hong Wang,Lin Luo,Zhili Xiao

A novel and efficient immunoaffinity column (IAC) based on bispecific monoclonal antibody (BsMAb) recognizing aflatoxin B1(AFB1) and ochratoxin A (OTA) was prepared and applied in simultaneous extraction of AFB1and OTA from food samples and detection of AFB1/OTA combined with ic-ELISA (indirect competitive ELISA). Two deficient cell lines, hypoxanthine guanine phosphoribosyl-transferase (HGPRT) deficient anti-AFB1hybridoma cell line and thymidine kinase (TK) deficient anti-OTA hybridoma cell line, were fused to generate a hybrid-hybridoma producing BsMAb against AFB1and OTA. The subtype of the BsMAb was IgG1via mouse antibody isotyping kit test. The purity and molecular weight of BsMAb were confirmed by SDS-PAGE method. The cross-reaction rate with AFB2was 37%, with AFG115%, with AFM148%, with AFM210%, and with OTB 36%. Negligible cross-reaction was observed with other tested compounds. The affinity constant (Ka) was determined by ELISA. The Ka (AFB1) and Ka (OTA) was 2.43 × 108L/mol and 1.57 × 108L/mol, respectively. Then the anti-AFB1/OTA BsMAb was coupled with CNBr-Sepharose, and an AFB1/OTA IAC was prepared. The coupling time and elution conditions of IAC were optimized. The coupling time was 1 h with 90% coupling rate, the eluent was methanol–water (60:40,v:v, pH 2.3) containing 1 mol/L NaCl, and the eluent volume was 4 mL. The column capacities of AFB1and OTA were 165.0 ng and 171.3 ng, respectively. After seven times of repeated use, the preservation rates of column capacity for AFB1and OTA were 69.3% and 68.0%, respectively. The ic-ELISA for AFB1and OTA were applied combined with IAC. The IC50(50% inhibiting concentration) of AFB1was 0.027 ng/mL, the limit of detection (LOD) was 0.004 ng/mL (0.032 µg/kg), and the linear range was 0.006 ng/mL~0.119 ng/mL. The IC50of OTA was 0.878 ng/mL, the LOD was 0.126 ng/mL (1.008 µg/kg), and the linear range was 0.259 ng/mL~6.178 ng/mL. Under optimum conditions, corn and wheat samples were pretreated with AFB1-OTA IAC. The recovery rates of AFB1and OTA were 95.4%~105.0% with ic-ELISA, and the correlations between the detection results and LC-MS were above 0.9. The developed IAC combined with ic-ELISA is reliable and could be applied to the detection of AFB1and OTA in grains.Keywords:aflatoxin B1;ochratoxin A;bispecific monoclonal antibody;immunoaffinity column;ic-ELISA

Development of an indirect competitive enzyme-linked immunosorbent assay for propiconazole based on monoclonal antibody

Jiao Li,Yuan Ding,He Chen,Wanlin Sun,Yue Huang,Fengquan Liu,Minghua Wang,Xiude Hua

Propiconazole is a widely used fungicide in agricultural production, but it poses risks to human health. Here, an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) based on a monoclonal antibody (mAb) was established to detect propiconazole residues. Three haptens of propiconazole were designed and synthesized to prepare antigens. Ten mAbs were obtained using hapten B (5-(4-(2-((1H-1,2,4-triazol-1-yl)methyl)-4-propyl-1,3-dioxolan-2-yl)-3-chlorophenyl) pentanoic acid). The ic-ELISA based on the combination of mAb 2G2E12 and hapten B-ovalbumin (coating antigen) showed the highest sensitivity. After optimization, the 50% inhibition concentration, limit of detection and linear range of the ic-ELISA were 2.33 μg/L, 0.26 μg/L and 0.26–21.28 μg/L, respectively. The cross-reactivities for the analogs of propiconazole were ≤1.9%. The average recoveries of the spiked samples were 75.7%–117.0% with relative standard deviations of ≤9.4%. The results of the ic-ELISA were consistent with those of gas chromatography-mass spectrometry in the analysis of blind samples. Therefore, we provide an alternative tool for the accurate and reliable determination of propiconazole.

Rapid screening of imidacloprid residue in grains and medicinal herbs: A newly designed hapten and monoclonal antibody

Yudan Wang,Jia’ an Qin,Jing Zhang,Ziyue Jin,Jiaoyang Luo,Meihua Yang

Three different imidacloprid hapten structures were designed to conjugate with proteins (bovine serum albumin, BSA; ovalbumin , OVA; keyhole limpet hemocyanin , KLH) for screening the optimal immunogen and coating antigen. Among these, an unreported antigen (hapten 6-KLH) was selected as the optimal immunogen and coating antigen. In addition, an imidacloprid-specific and high titer monoclonal antibody (IMIB7C3) was obtained by using the above-selected immunogen. A sensitive ic-ELISA (indirect competitive enzyme-linked immunosorbent assay) with a half-maximal inhibitory concentration (IC 50 ) of 1.3 ng mL −1 was established by using the IMIB7C3 antibody (only 1.2 ng per well) to detect the residues of imidacloprid in grains (wheat and maize) and different herbs ( Notoginseng radix et rhizoma, Dioscoreae rhizoma, Lonicerae japonicae flos, Astragali radix , Jujubae fructus ). The detection results of real samples by the developed immunoassay were confirmed by liquid chromatography-tandem mass spectrometry (LC–MS/MS), which proved the accuracy and reliability of the established ic-ELISA. These results indicate that the proposed ic-ELISA method is suitable for rapid and high-throughput detection of imidacloprid residues in agricultural products and medicinal herbs. Furthermore, a quantitative risk assessment was conducted for Lonicerae japonicae flos based on the detection results, which indicates an acceptable risk to human health after the intake of Lonicerae japonicae flos polluted by imidacloprid.

Development of broad-specific monoclonal antibody-based immunoassays for simultaneous ochratoxin screening in medicinal and edible herbs

Mengyue Guo,Jing zhang,Jianxin Lv,Tongwei Ke,Jiao Tian,Kun Miao,Yudan Wang,Dandan Kong,Haonan Ruan,Jiaoyang Luo,Meihua Yang

The frequent occurrence of ochratoxins in food and herbs poses a considerable threat to consumer and animal health. Here, we generated a broad-specificity monoclonal antibody against three ochratoxins. Concentrations for 50% inhibition (IC 50 ) for OTA, OTB, and OTC were 0.37, 0.23, and 2.24 ng/mL, respectively. In the two matrices, the proposed indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) exhibited a limit of detection (LOD) of 1.5 μg/kg for OTA and 1.0 μg/kg for OTB, respectively, whereas the cut-off levels of lateral flow immunoassay (LFIA) for OTA and OTB were respectively determined to be 1 and 0.25 ng/mL. As for OTC, the LOD of ic-ELISA and cut-off concentration of LFIA were 15.5 μg/kg and 8 ng/mL for milkvetch root and 37.5 μg/kg and 16 ng/mL for ginger, respectively. The real sample detection results by ic-ELISA and LFIA methods were validated via LC-MS/MS analysis. Therefore, the developed immunoassays are useful and effective tools for high-throughput screening and rapid on-site determination of ochratoxins in complex matrices.

Quantitative determination of prothioconazole in wheat grain, soybean, and pond water based on a polyclonal antibody

Zhihui Guo,Mingya Deng,Wenwen Fang,Kang Zheng,Min Liao,Yulong Wang,Qingkui Fang

Prothioconazole and its metabolite are considered a potential threat to human health and environmental safety. Thus, the development of a sensitive and rapid detection method for prothioconazole is crucial to ensure the safety of agricultural products. In this study, a new hapten of prothioconazole was designed and synthesized, and a selective polyclonal antibody with high affinity against prothioconazole was produced, which was obtained from immunized New Zealand white rabbits. Based on the polyclonal antibody, an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and indirect competitive chemiluminescence enzyme immunoassay (ic-CLEIA) were developed for detecting prothioconazole pesticides. Under optimized experimental conditions, the limit of quantification (LOQ) values for ic-CLEIA and ic-ELISA were 1.8 and 10.7 ng mL−1, respectively. The results demonstrated that the sensitivity (LOQ) achieved by ic-CLEIA was more than five times higher compared to that obtained with ic-ELISA. In addition, the recoveries obtained by adding standard prothioconazole to wheat grain, soybean, and pond water samples were in the range of 81.9 to 104.7% for ic-ELISA and 89.0 to 118.0% for ic-CLEIA.