Analytical Methods
·
2014
·Vol. 6
·期 7
·2306-2313
·
IF 1.938
Jianhua Shu,Liming He,Huanzhong Ding,Liqi Wang,Hongyan Guo,Yan Gao,Emmanuel E. Dzakah,Zhenling Zeng
A sensitive and specific monoclonal enzyme-linked immunosorbent assay (ELISA) for the determination of tissue-bound metabolite 3-amino-5-morpholinomethyl-2-oxazolidone (AMOZ) was developed. Three different haptens of AMOZ were synthesized. The haptens (I, III) were derived from 4-carboxybenzaldehyde and ethyl 4-bromobutyrate, respectively. Hapten II was based on hapten I but a CN bond in the parent structure was reduced. Corresponding immunogens and coating antigens were prepared to improve the assay sensitivity. The hybridomas 4 × 108 secreting antibodies against CPAMOZ were obtained from immunogens I-BSA by monoclonal antibody (mAb) technology. The antibody indicated that: (1) the small molecular AMOZ cannot induce the specific response against AMOZ; (2) the weakened conjugation effect improved the recognition of AMOZ. Assessment of twelve coating antigen/antibody combinations consequently resulted in the development of an indirect competitive enzyme-linked immunosorbent assay (icELISA). The results showed that the sensitivity was highly improved about fifteen-fold for the novel heterologous coating antigen at low hapten density. The IC50 value of the ELISA for CPAMOZ was 0.13 ng mL−1. The cross reactivity values of the assay with NPAMOZ, AMOZ and FTD were 118%, 2.3% and 16.3%. Recoveries from AMOZ fortified animal tissues were in a range of 82.6–108.4% and the CV values were less than 12.5%. The immunoassay was further validated by a LC-MS/MS method and the two methods showed good correlation (r2 = 0.9908). The proposed icELISA could be used for an accurate monitoring of AMOZ in animal tissues.
研究领域:
分析化学
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免疫学
·
兽医学
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高影响因子
Scientific Reports
·
2024
·Vol. 14
·期 1
·1-11
·
IF 3.8
Zheng Hailing,Yang Ruling,Guo Jianbo,Xie Zhenbin,Liu Jingxuan,Yang Hailiang,Jia Liling,Yu Jian,Cai Qiutong,Tang Fei,Zhou Yang
Silk was the driving force behind the opening of the Silk Road, positioning this material as a global commodity that greatly influenced the progress of human civilization. Due to the natural protein properties of silk, the internal structure and external characteristics of silk cultural relics are irreversibly destroyed during the process of burial or when passed down through the generations until the production information and material components completely disappear, making it difficult to obtain direct archaeological evidence for pinpointing the origin of silk. The sacrificial pits at the Sanxingdui archaeologcal site, located in Guanghan, Sichuan, China, have been found with layers of ash above the layers of artifacts and some bronzes with fabric traces. Among the artifacts, one grid-like ware artifact first appeared in the Bronze Age in China. The two sides of the grid-like ware were grid-like ovals made of bronze material, and inside, there was an oval-shaped complete piece of jade with a bent back. Fabric traces were found on both the jade and bronze surfaces. In order to determine the specific function of fabric at this site, the developed silk fibroin immunoaffinity column (IAC) enrichment technique combined with enzyme-linked immunosorbent assay (ELISA), morphology observation and proteomics were used to identify mineralized fabric material and fabric residues in the ash layer. Silk residues were successfully detected, which confirmed the early use of silk as a material carrier to communicate between Heaven and Earth and provided archaeological evidence for the cultural origins of silk.
研究领域:
考古学
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材料科学
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文化遗产研究
·
蛋白质组学
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DOI
PubMed
高影响因子
FOOD RESEARCH INTERNATIONAL
·
2026
·
IF 8.8
Yuanyuan Zhao,Yongmao Lu,Tian Yang,Shaohua Hu,Yuanyuan Yan,Xingguo Wang,Wei Wei
Multi-method strategy enhances accuracy in human milk vitamin research. • Mass spectrometry achieved precise 25(OH)D₂/₃ quantification in human milk. • Immunoassays offered rapid screening but showed a 72.98% positive bias. • Liquid chromatography profiled vitamins A and E but lacked sensitivity for D and K. • 25(OH)D₃ increased slightly, while 25(OH)D₂ declined across lactational stages. Accurate analysis of vitamin D in human milk is essential for infant nutrition but remains analytically challenging due to low levels and a complex matrix. This study evaluates a practical multi-method strategy using LC-MS/MS, ELISA, and UPLC-PDA for vitamin D analysis in human milk. LC-MS/MS precisely quantified 25(OH)D₂ and 25(OH)D₃ with assay LOQ of 0.045 ng/mL and 0.100 ng/mL, respectively. ELISA showed a 11.58 ng/mL positive bias of vitamin D which limits use for accurate quantification. UPLC-PDA profiled vitamins A and E effectively but lacked sensitivity for vitamin D. Longitudinally, LC-MS/MS results show 25(OH)D₃ and total 25(OH)D concentrations remained relatively stable, with medians ranging from 0.141 to 0.281 ng/mL and 0.263 to 0.315 ng/mL, respectively, across the study period, while 25(OH)D₂ peaked at 8 weeks before declining. This study establishes LC-MS/MS as the preferred analytical strategy for quantifying 25(OH)D 2 and 25(OH)D 3 in human milk, providing critical data to assess infant nutritional intake and guide clinical interventions. Download: Download high-res image (160KB) Download: Download full-size image
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MOLECULES
·
2023
·Vol. 28
·期 15
·5743
·
IF 4.6
Yexuan Ren,Ruwen Tian,Ting Wang,Junlin Cao,Jianguo Li,Anping Deng
In this study, an extremely highly sensitive enzyme-linked immunosorbent assay (ELISA) based on a newly produced monoclonal antibody (mAb) for the detection of ochratoxin A (OTA) in food samples was developed. OTA-Bovine serum albumin (BSA) conjugate was prepared and used as the immunogen for the production of the mAb. Among four hybridoma clones (8B10, 5C2, 9B7, and 5E11), the antibody from 8B10 displayed the highest affinity recognition for OTA. Based on the mAb (8B10), the IC50and LOD of the ELISA for OTA were 34.8 pg mL−1and 1.5 pg mL−1, respectively, which was 1.53~147 times lower than those in published ELISAs, indicating the ultra-sensitivity of our assay. There was no cross-reactivity of the mAb with the other four mycotoxins (AFB1, ZEN, DON, and T-2). Due to the high similarity in molecular structures among OTA, ochratoxin B (OTB), and ochratoxin C (OTC), the CR values of the mAb with OTB and OTC were 96.67% and 22.02%, respectively. Taking this advantage, the ELISA may be able to evaluate total ochratoxin levels in food samples. The recoveries of the ELISA for OTA in spiked samples (corn, wheat, and feed) were 96.5–110.8%, 89.5–94.4%, and 91.8–113.3%; and the RSDs were 5.2–13.6%, 8.2–13.0%, and 7.7–13.7% (n= 3), respectively. The spiked food samples (corn) were measured by ELISA and HPLC-FLD simultaneously. A good correlation between ELISA (x) and HPLC-FLD (y) with the linear regression equation of y = 0.918x − 0.034 (R2= 0.985,n= 5) was obtained. These results demonstrated that the newly produced mAb-based ELISA was a feasible and ultra-sensitive analytical method for the detection of OTA in food samples.Keywords:enzyme-linked immunosorbent assay (ELISA);monoclonal antibody (mAb);mycotoxin;ochratoxin A (OTA);food samples;sensitive detection
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PubMed
高影响因子
SCIENCE OF THE TOTAL ENVIRONMENT
·
2021
·
IF 7.963
Nuanfei Zhu,Xuesong Li,Ye Liu,Jingfu Liu,Yawei Wang,Xiangyang Wu,Zhen Zhang
Dibutyl phthalate (DBP) is considered as one of the most widely used phthalate esters (PAEs), which has attracted worldwide concerns because of its potential threats to eco-environments and human health. Systematic investigations of DBP environmental occurrence contribute to the further risk assessment, which depends on effective and available analytical methods. In this study, an amplified ratiometric fluorescence ELISA was established for sensitive and high-throughput detection of DBP in the aquatic system based on a novel tetrahedral DNA nanostructure (TDN)-scaffolded-DNAzyme (Tetrazyme). Wherein, Tetrazyme was prepared by the precise folding of G-quadruplex sequence on three vertex angles of the TDN, together with hemin as the horseradish peroxidase (HRP)-mimicking enzyme. The rigid TDN avoided the local overcrowding effect to provide a reasonable spatial spacing on the interface for G-quadruplex sequence, increasing the collision chance between DNAzyme and substrates, improving the catalytic ability of DNAzyme effectively. Besides, streptavidin (SA) and biotin (bio) were used to anchor TDN and antibody, in which the specific binding of SA/bio could make more Tetrazyme conjugate on each signal element, resulting in the dual signal amplification. Meanwhile, the accuracy and precision were enhanced owing to the inherent built-in rectification to the environment from the dual output ratiometric fluorescence assay. Under the optimized conditions, the detection limit of this proposed method was 0.17 ng/mL (16 times lower than that of conventional ELISA using the same antibody) with a satisfactory accuracy (recoveries, 79.0%- 116.2%; CV, 2.1–6.5%). Overall, this platform provides a promising way for accurate, sensitive and rapid determination of DBP from environmental waters.
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PubMed
高影响因子
TALANTA
·
2022
·
IF 6.556
Ting Zhou,Guangzhao He,Changjia Hu,Kang Wu,Ying Liu,Jianguo Li,Anping Deng
5-Fluorouracil (5-FU) is an effective anticancer drug widely used in cancer treatment. In this study, two 5-FU derivatives containing a spacer arm with the carboxylic group at the end were synthesized, which were linked to the carrier proteins to form 5-FU-protein conjugates used as the immunogens for the production of monoclonal antibody (mAb). Based on the produced mAb, the highly sensitive and specific enzyme-linked immunosorbent assay (ELISA) for 5-FU detection was established. The IC 50 and LOD values of the assay were found to be 19.5 ng mL −1 and 0.5 ng mL −1 , respectively. There was no cross-reactivity (CR) of the ELISA with cytosine , thymine and uracil , which avoided the interference from inherent pyrimidines. The CR values of the assay with three substitutes of 5-FU (tegafur, 5-fluoro-2′-deoxyuridine, carmofur) were within 9.7%–17.6%. The produced mAb was also applied in sample extraction. The immuno-affinity column capable specific capturing 5-FU was prepared by immobilizing the mAb on Sepharose-4B gel and filling into a SPE column. The recoveries of 5-FU in spiked samples measured by ELISA were 72.4%–90.7% with RSD of 3.6%–8.3%. Five blood samples collected from patients were extracted by immuno-affinity column, then measured by ELISA and confirmed by HPLC-MS/MS. There was a good correlation between HPLC-MS/MS and ELISA. It is demonstrated that the developed ELISA combined with immuno-affinity extraction can be a powerful alternative method for the detection of 5-FU in blood samples.
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高影响因子
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
·
2025
·
IF 8.5
Junru Sun,Ruiqin Zhu,Mengxiang Wang,Jinxing Song,Lei Zhou,Zhuoya Sun,Yanze Li,Liuyang Jiao,Lu Xia,Hua He,Gaiping Zhang,Yanan Wu
Porcine epidemic diarrhea (PED) causes significant economic losses to the pig farming industry worldwide and currently lacks an effective vaccine. Multiple detection assays and protein purification methods use magnetic nanoparticles due to their biocompatibility, high specific surface area, and solution suspension properties. In this study, a two-probe competitive ELISA based on magnetic nanoparticles for detecting PEDV N protein was developed. MNPs-N and McAb-HRP probes were prepared and the procedure was optimized to identify the ideal reaction conditions. Compared to other methods, the developed method shortens the detection time to 50 min. The coefficient of variation (CV) for intra- and inter-lot replicates was less than 10 %, with reproducibility. The coincidence rate with commercial kits is 93.07 %, making this method reliable and suitable for PED immune monitoring and diagnostics.
研究领域:
兽医学
·
免疫学
·
分子生物学
·
纳米技术
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DOI
PubMed
高影响因子
Analytical Methods
·
2024
·
IF 2.7
Gaoshuang Hu,Ziyang Liu,Yiqin Zhang,Tao Zhao,Zhuqing Xing,Shan Gao,Jianxiong Hao
Long-term consumption of foods with excessive enrofloxacin (ENRO) residues may cause the accumulation of ENRO in the human body, thus damaging human health. In this study, quantum dot-based biomimetic fluorescence immunoassays were used for enrofloxacin detection in food of animal origin. Under the most suitable conditions, the detection limit (IC15) of the method in standard solution was 0.13 ± 0.02 ng L−1 and the sensitivity (IC50) was 0.13 ± 0.18 μg L−1. The recoveries for ENRO from four fortified food samples, including chicken, eggs, shrimp, and milk, ranged from 85.74% to 114.19% and the coefficients of variation were 0.01–18.09%. The established method shows good agreement with the results obtained using commercial ELISA kits, with a correlation coefficient of 0.997. The proposed method shows the advantages of high sensitivity, specificity and wide detection range. It can be used as an alternative method for the rapid and sensitive detection of ENRO in food of animal origin.
研究领域:
食品科学
·
分析化学
·
生物技术
·
兽医学
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DOI
PubMed
高影响因子
JOURNAL OF FOOD COMPOSITION AND ANALYSIS
·
2025
·
IF 4.6
Wenjing Liu,Guangjie Wu,Liping Wang,Jiaying Zhao,Renrong Liu
Immunoaffinity column (IAC) cleanup, coupled with high performance liquid chromatography (HPLC) analysis, is one of the most reliable methods for aflatoxin B1 (AFB1) analysis. However, real-time monitoring of the IAC capacity remains unachievable in IAC-HPLC analysis, compromising the reliability of AFB₁ quantification. Two artificial structural analogues of aflatoxin (AFB1-OH and AFB2-OH) were synthesized as potential markers for IAC capacity monitoring. Competitive ELISA and recovery experiments demonstrated that AFB2-OH was more suitable as a marker. When AFB2-OH was used for real-time monitoring of the capacity of the IAC in AFB1 determination by HPLC, the recovery of AFB1 exceeded 95 %. The limit of detection (LOD) and the limit of quantification (LOQ) for AFB1 were determined to be 0.14 ng/mL and 0.46 ng/mL, respectively. The method also exhibited good precision and accuracy. In spiked recovery experiments of four grains, the recovery of AFB1 ranged from 85.16 % to 107.05 % with relative standard deviations of 0.89 %-4.98 %. More importantly, due to the addition of AFB2-OH, the capacity of the IAC was monitored in real-time, ensuring the reliability of AFB1 determination results.
研究领域:
分析化学
·
食品安全
·
生物化学
·
色谱分析
·
免疫分析
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高影响因子
JOURNAL OF DAIRY SCIENCE
·
2020
·
IF 3.333
Suhua Wang,Bolong Fang,Meifang Yuan,Zexiang Wang,Juan Peng,Weihua Lai
ABSTRACT In this study, a novel colorimetric and fluorescent dual-mode ELISA based on glucose oxidase (GOx)-triggered Fenton reaction was developed for the qualitative and quantitative detection of danofloxacin (DAN). In this system, streptavidin-linked biotinylated anti-DAN-monoclonal antibody (SA-Bio-mAb) and biotinylated GOx (Bio-GOx) form the immune complex mAb-Bio-SA-Bio-GOx. In the absence of DAN, the mAb-Bio-SA-Bio-GOx would be immobilized by combining with coated DAN-BSA and catalyzed glucose to generate H 2 O 2 . The Fenton reaction between H 2 O 2 and Fe 2+ generated hydroxyl radicals, which oxidized the o -phenylenediamine to 2,3-diamino-phenazine. A dual-signal immunoassay with colorimetry and fluorescence as the signal readout was established. In the presence of DAN, DAN and DAN-BSA competed with Bio-mAb, decreasing the connection between immune complexes and DAN-BSA and finally resulting in lower signal of colorimetry and fluorescence. Under optimal conditions, the limit of detection of the fluorescence immunoassay was 0.337 ng/mL and was 5.24-fold lower than that of traditional ELISA. The colorimetric immunoassay cut-off value was 30 ng/mL in milk. The average recoveries of the method for milk samples that are spiked with different concentrations of DAN were 91.1 to 128.3%, with a coefficient of variation of 0.7 to 8.2%. These results of the method exhibited good agreement with those of liquid chromatography-tandem mass spectrometry system (LC-MS/MS) method. In brief, this work provides an improved screening strategy with high sensitivity and accuracy for the qualitative or quantitative detection of DAN in milk monitoring.
研究领域:
分析化学
·
食品安全
·
生物化学
·
兽医学
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DOI
PubMed
高影响因子
BIOSENSORS & BIOELECTRONICS
·
2020
·
IF 10.257
Shengfeng Huang,Rentao Tang,Tingting Zhang,Jing Zhao,Zhengjin Jiang,Qiqin Wang
In this study, a high-performance anti-fouling coating based on poly adenine (polyA n ) as well as a highly specific cluster of differentiation 20 (CD20) epitope mimetic peptide (CN14) were employed to synergistically construct a facile biosensor for the rapid and sensitive determination of rituximab in lymphoma patients' plasma. The well-designed and optimized polyA n coating displayed excellent stability, hydrophilicity , thanks to its intrinsic affinity with gold surface and thoroughly exposed hydrophilic phosphate groups. Moreover, the proposed strategy avoids the necessity to modify binding groups (e.g. thiol), making it more facile, repeatable and efficient. When dealing with complex clinical plasma samples, the polyA n coating demonstrated better anti-fouling performance and lower background signal in comparison with mercaptan and bovine serum albumin coatings. The dissociation constant (~60 nM) between CN14 and rituximab was measured by microscale thermophoresis and their binding mechanism was further explained using computer simulation. The constructed GE/CN14/polyA 20 biosensor displayed satisfactory performance with detection limit of 35.26 ng/mL. Finally, the proposed biosensor was successfully applied for rapidly determining rituximab in lymphoma patients' plasma, and exhibited comparable accuracy to the commercial ELISA , but has advantages including a shorter detection time, wider detection range and lower cost. It's worth noting that the anti-fouling polyA n coating can be tailored according to the surface property of sensing interface and can be easily expanded to other gold electrode related biosensors.
研究领域:
生物医学工程
·
临床化学
·
生物传感器
·
癌症研究
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PubMed
高影响因子
FOOD CHEMISTRY
·
2024
·
IF 8.5
Bolong Fang,Xin Liu,Juan Peng,Yuzhi Li,Zhiyong Gong,Weihua Lai
In this study, a type of luminescent porous coordination network-224 (PCN-224) in alkaline conditions was synthesized with the dramatic fluorescence enhancement by 20.4 times, which was explained by the fact that the decrease of Zr 4+ content in alkaline conditions resulted in the partial recovery of the electron cloud density of 4,4′,4′′,4′′′-(Porphine-5,10,15,20-tetrayl) tetrakis(benzoic acid) (TCPP). Given the large overlap between the excitation spectrum of PCN-224 and the absorption band of Ag nanoparticles (Ag NPs), the coating of the Ag layer on PCN-224 triggered the fluorescence quenching effect, which was applied to “turn off” fluorescence immunoassay for sensitive detection of Escherichia coli O157:H7 ( E. coli O157:H7) in milk. The proposed immunoassay reached a low limit of detection (LOD) of 3.3 × 10 2 CFU mL −1 , 29.7 times more sensitive than the conventional ELISA. It will provide a novel alternative strategy for sensitively detecting pathogenic bacteria in the field of food safety.
研究领域:
材料科学
·
分析化学
·
食品安全
·
生物传感器
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PubMed
高影响因子
危险品化学品经营许可证（带存储）