BIOSENSORS & BIOELECTRONICS
·
2025
·
IF 10.5
Junjie Zeng,Yanping Li,Qinghua He,Ji Xia,Jiebiao Guo,Zhui Tu
Phage display-mediated immuno-polymerase chain reaction (PD-IPCR) combines the strengths of phage display technology and immuno-PCR. The link between genotype and phenotype has been established by displaying antibody fragments on the surface of phage particles, allowing signal transmission. In this study, we utilized aflatoxin B 1 (AFB 1 ) as a model system to investigate the efficacy of PD-IPCR. Signal amplification was achieved using both real-time fluorescence quantitative PCR (qPCR) and a custom-made droplet digital PCR (dPCR) system. Under optimized conditions, the assay using dPCR achieved a limit of detection of 0.015 ng/mL, a 6-fold improvement over qPCR. The validation of our method using major agricultural produce, such as corn, flour, and soy sauce, shows excellent recoveries ranging from 85.77 % to 108.28 %, with a total assay time of less than 2 h. Furthermore, we compared our results with a high-performance liquid chromatography (HPLC) method for AFB 1 content in corn and flour, revealing consistent outcomes (r = 0.9847). These results demonstrate the potential of nanobody-based PD-IPCR as a reliable tool for detecting mycotoxins.
研究领域:
食品安全
·
分子生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
ACS Applied Materials & Interfaces
·
2016
·
IF 7.145
Guanghui Li,Min Zhu,Lu Ma,Junrong Yan,Xiaoling Lu,Yanfei Shen,Yakun Wan
A phage display library of variable domain of the heavy chain only antibody or nanobody (Nb) was constructed after immunizing a bactrian camel with testosterone. With the smaller molecular size (15 kDa), improved solubility, good stability, high affinity, specificity, and lower immunogenicity, Nbs are a promising tool in the next generation of diagnosis and medical applications. Testosterone is a reproductive hormone, playing an important role in normal cardiac function and being the highly predictive marker for many diseases. Herein, a simple and sensitive immunosensor based on electrochemical impedance spectroscopy (EIS) and Nbs was successfully developed for the determination of testosterone. We successfully isolated the antitestosterone Nbs from an immune phage display library. Moreover, one of the Nbs was biotinylated according to in vivo BirA system, which showed the highest production yield and the most stable case. Further, the EIS immunosensor was set up for testosterone detection by applying the biotinylated antitestosterone Nb. As a result, the biosensor exhibited a linear working range from 0.05 to 5 ng mL–1 with a detection limit of 0.045 ng mL–1. In addition, the proposed immunosensor was successfully applied in determining testosterone in serum samples. In conclusion, the proposed immunosensor revealed high specificity of testosterone detection and showed as a potential approach for sensitive and accurate diagnosis of testosterone.
研究领域:
生物技术
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医学诊断
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分析化学
✓ 已验证
DOI
PubMed
高影响因子
FOOD CHEMISTRY
·
2025
·
IF 9.8
Wei Zeng,Mingrui Lu,Long Wu,Xin Gao,Qi Chen,Sihang Zhang
A Raman scattering and nanozyme-linked immunosorbent assay (NLISA) dual modal immunosensor, was constructed by mesoporous SiO 2 /Au-Pt nanozymes (m-SAP) and nanobodies (A2.3-SBP). Oxidized TMB served as Raman and ELISA signals in a competitive binding assay. Under optimized conditions, an inverse correlation was established between the Microcystin-LR (MC-LR) concentration and the signals, spanning Raman and ELISA ranges of 0.1–100 μg L −1 and 1.0–500 μg L −1 , with limit of detections (LODs, 3σ/S) of 0.015 μg L −1 and 0.12 μg L −1 , respectively. The LODs showed over 90 times and 11 times higher sensitivity than that of traditional ELISA (t-ELISA, LOD, 1.36 μg L −1 ). The immunosensor exhibited excellent accuracy in practical samples, can be integrated together for the detection of MC-LR within 45 min, which greatly short the detection time of t-ELISA (>2 h). This method displayed potential for detecting other toxins by simply changing the nanobodies.
研究领域:
生物化学
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环境科学
·
分析化学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
ACS Applied Materials & Interfaces
·
2025
·
IF 8.2
Yiqi Zeng,Siyu Zhou,Yicheng Yang,Bin Tang,Weijun Wei,Gang Huang,Changfeng Wu,Xiaofeng Fang
Nanobodies have gained significant attention as promising tools for cancer diagnostics and treatment due to their unique ability to precisely target specific cancer cells. However, a major challenge lies in the site-specific incorporation of multifunctional molecules into nanobodies, as it is essential to link these molecules in a manner that preserves the nanobody’s function and stability while retaining the desired therapeutic or diagnostic properties. This study outlines the development of dual-functional nanobody optical probes for enhanced cancer diagnostics and therapeutic interventions. We designed a dual-functional clickable linker that enables site-specific functionalization of the nanobody, facilitating the simultaneous conjugation of two dyes: indocyanine green for imaging and chlorin e6 for photodynamic therapy. In vitro cellular assays confirmed the successful labeling of the dual-functional dyes, with the nanobody probe exhibiting high cellular binding specificity. In vivo imaging of mice bearing Hep3B tumors revealed clear visualization with a high signal-to-noise ratio. Furthermore, PEGylated probes significantly improved tumor retention, enhancing both imaging contrast and photodynamic therapy efficacy as compared to free chlorin e6. These dual-functional nanobody probes show great promise for the precise diagnosis and treatment of malignant tumors.
研究领域:
生物医学工程
·
肿瘤学
·
分子影像学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
Toxins
·
2022
·Vol. 14
·期 12
·821
·
IF 5.075
Yi Chen,Guanggang Qu,Hongkun Quan,Yihui Wang,Changjiang Wang,Md Atiqul Haque,Cheng He
Background: Fumonisin B1 (FB1) is a secondary metabolite produced mainly byFusarium verticillioidesorFusarium proliferatum. It poses a huge threat to the sustainable animal industry and human health as well via food chains (egg, meat and milk). AlthoughE. coli-expressed nanobodies are documented for diagnostic applications, nanobodies remain elusive as FB1 detoxifiers in feed and food. Results: In the present study, theE. coli-expressed nanobody was assessed to remove FB1 in fresh milk, embryonated eggs and broilers. Firstly, 2 alpacas received intramuscularly FB1-adjuvanted BSA 6 times, and then the variable domain of the heavy-chain antibody (VHH) of fb1 genes were amplified to clone into the pCANTAB 5 E vector in order to generate a VHH-FB1 phage antibody display library, yielding 3.4 × 1010capacity with 96.7% positivity. Afterwards, 5 anti-FB1 nanobodies were expressed and identified. Furthermore, maximal 43.2% FB1 was removed from milk by 1:2000 concentration of nanobody 5 (Nb5). Furthermore, SPF-embryonated eggs were inoculated into albumens with nanobody-treated FB1. The Nb5 group yielded an 83.3% hatching rate, higher body weight, lower gizzard ulceration and fewer FB1 residuals. In order to warrant the above results, 50 broilers aged 10 days were received orally with 20 ppm of FB1 for 20 days. At the same time, birds were fed orally with 50 μg of Nb5 or bivalent nanobody 11 (BiNb11). Finally, the Nb5 group showed a higher relative body weight gain and lower gastric ulcerations and fewer inflammations in the thymus and bursa. Conclusions: Based on the above evidence, the Nb5 nanobody may be considered as an additional FB1 detoxifier, contributing to FB1 decontamination.Keywords:fumonisin B1;nanobody;phage-display technology;detoxification;efficacy testKey Contributions:1. Five nanobodies were firstly expressed and bound specifically to FB1 mycotoxin. 2. Monomer nanobody Nb5 is a promising food detoxifier by degrading FB1-contaminated fresh milk. 3. Engineered nanobody Nb5 is a potential feed detoxifier for poultry industry.
研究领域:
食品科学
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毒理学
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动物健康
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生物技术
✓ 已验证
产品关联
DOI
PubMed
高影响因子
RSC Advances
·
2015
·Vol. 5
·期 94
·77380-77387
·
IF 3.84
Ying Xiong,Zhui Tu,Xiaolin Huang,Bing Xie,Yonghua Xiong,Yang Xu
The magnetic-bead-based immunoaffinity extraction (M-IAE) method has been widely used for the purification of aflatoxin B1 (AFB1) from complex food and feed matrices. However, this method suffers from several inherent disadvantages that limit its practical application, including low reusability and limited saturated adsorption capacity. Herein, we report an improved M-IAE for the highly efficient purification of AFB1 from corn samples. This method involves the expression of anti-AFB1 nanobodies (Nbs), which possess high degeneration resistance, to replace conventional antibodies; and magnetic beads carrying poly(acrylic acid) brushes (MB@PAA) were fabricated as an “Nb container” for improving AFB1 adsorption capacity. The MB@PAA shows a high loading capacity for anti-AFB1 Nbs at 623 μg g−1, which is 19-fold the magnitude of that of conventional MB@Nbs. Meanwhile, the resultant MB@PAA@Nbs exhibit good AFB1 adsorption, with a maximum adsorption capacity of 0.23 mg g−1, which is 35-fold superior to that of the conventional MB@Nbs. MB@PAA@Nbs can be reused at least 10 times, without obvious loss of the capture efficiency for AFB1. The reliability and practicability of the proposed MB@PAA@Nbs for AFB1 extraction were further evaluated using AFB1-spiked corn samples. In brief, the proposed MB@PAA@Nbs-based immunoaffinity extraction method is a highly promising, novel sample pre-treatment platform for AFB1 as well as other mycotoxins.
研究领域:
食品安全
·
分析化学
·
免疫学
✓ 已验证
DOI
高影响因子
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
·
2021
·
IF 6.953
Ting He,Yao Nie,Tingting Yan,Jiang Zhu,Xiaoling He,Ying Li,Qi Zhang,Xiaoqian Tang,Rui Hu,Yunhuang Yang,Maili Liu
Nanobodies (Nbs) have shown great potential in immunodetection of small-molecule contaminants in food and environmental monitoring. However, the limited knowledge of the mechanism of Nbs binding to small molecules has hampered the development of high-affinity Nbs and assay improvement. We previously reported two homologous nanobodies Nb26 and Nb28 specific to aflatoxin B 1 (AFB 1 ), with the former exhibiting higher sensitivity in ELISA. Herein, Nb26 was selected as the model antibody to resolve its solution nuclear magnetic resonance (NMR) structure, and investigate its AFB 1 recognition mechanism. The results revealed that Nb26 exhibits a typical immunoglobulin fold and its AFB 1 -binding interface is uniquely located in complementarity-determining region 3 (CDR3) and framework region 2 (FR2). This finding was applied to improve the binding activity of Nb28 against AFB 1 by constructing two Nb28-based mutants A 50 V and S 102 D, resulting in 2.3- and 3.3-fold sensitivity enhancement over the wild type, respectively. This study develops an NMR-based strategy to analyze the underlying mechanism of Nb against AFB 1 , and successfully generated two site-modified Nbs with improved detection sensitivity. It is believed that this work could greatly expand the applications of Nbs by providing a way to enhance the binding activity.
研究领域:
食品科学
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环境监测
·
生物化学
·
分子生物学
✓ 已验证
DOI
PubMed
高影响因子
Microbiology Spectrum
·
2021
·Vol. 9
·期 3
·
IF 7.171
Song Zhu,Fei Luo,Bin Zhu,Fei Ling,Er-Long Wang,Tian-Qiang Liu,Gao-Xue Wang
Viral diseases of the central nervous system (CNS) represent a major global health concern. Difficulties in treating these diseases are caused mainly by the biological tissues and barriers, which hinder the transport of drugs into the CNS. To counter this, a nanobody-mediated virus-targeting drug delivery platform (SWCNTs-P-A-Nb) is constructed for CNS viral disease therapy. Viral encephalopathy and retinopathy (VER), caused by nervous necrosis virus (NNV), is employed as a disease model. SWCNTs-P-A-Nb is successfully constructed by employing single-walled carbon nanotubes, amantadine, and NNV-specific nanobody (NNV-Nb) as the nanocarrier, anti-NNV drug, and targeting ligand, respectively. Results showed that SWCNTs-P-A-Nb has a good NNV-targeting ability in vitro and in vivo, improving the specific distribution of amantadine in NNV-infected sites under the guidance of NNV-Nb. SWCNTs-P-F-A-Nb can pass through the muscle and gill and be excreted by the kidney. SWCNTs-P-A-Nb can transport amantadine in a fast manner and prolong the action time, improving the anti-NNV activity of amantadine. Results so far have indicated that the nanobody-mediated NNV-targeting drug delivery platform is an effective method for VER therapy, providing new ideas and technologies for control of the CNS viral diseases.IMPORTANCE CNS viral diseases have resulted in many deadly epidemics throughout history and continue to pose one of the greatest threats to public health. Drug therapy remains challenging due to the complex structure and relative impermeability of the biological tissues and barriers. Therefore, development in the intelligent drug delivery platform is highly desired for CNS viral disease therapy. In the study, a nanobody-mediated virus-targeting drug delivery platform is constructed to explore the potential application of targeted therapy in CNS viral diseases. Our findings hold great promise for the application of targeted drug delivery in CNS viral disease therapy.
研究领域:
医学
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生物技术
·
药理学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
Small
·
2025
·
IF 12.1
Liyuan Zhao,Yujuan Cao,Yanru Xin,Chenxi Liu,Jin Yang,Yanan Li,Sidan Tian,Zhenxing Liu,Haibo Jia,Mugen Liu,Min Hu,Liang Luo,Fanling Meng
α-Synuclein (α-Syn) amyloidogenesis is considered a promising diagnostic marker and therapeutic target for Parkinson's disease (PD). Simultaneously visualizing and mitigating α-Syn amyloidogenesis are essential for future PD theranostics, yet they continue to pose an insurmountable challenge. This study have herein developed a nanobody-decorated polydiacetylene to approach a straightforward solution. Grafting α-Syn61-95 segment into the third complementary determining region of a parent nanobody generates an engineered nanobody X30 that can bind with α-Syn and prevent its amyloidogenesis through homotypic interaction. It next use X30 to decorate poly(deca-4,6-diynedioic acid) (PDDA), a polydiacetylene with an ultrastrong alkyne Raman signal (2120 cm −1 ) in the cellular silent region, to create an α-Syn targeting Raman probe PX30. The binding affinity between X30 and α-Syn can be further boosted for over 150 times attributed to the rigidity of PDDA backbone and the multivalent effect. Therefore, PX30 not only enables real-time Raman visualization of α-Syn amyloidogenesis with a high signal-to-noise ratio in living zebrafish, but also alleviates amyloidogenesis-mediated damage to zebrafish embryos by effectively inhibiting α-Syn amyloidogenesis at low stoichiometric concentrations and scavenging pathologic reactive oxygen species.
研究领域:
神经科学
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分子生物学
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药物开发
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生物医学工程
✓ 已验证
产品关联
DOI
PubMed
高影响因子
MICROCHIMICA ACTA
·
2024
·Vol. 191
·期 11
·1-12
·
IF 5.3
Wen Peng,Su Hao,Yin Wen-jia,Hu Jian-cheng,Wang Yu,Yang Jin-yi,Xiao Zhi-li,Xu Zhen-lin,Shen Yu-dong,Wang Hong,Hammock Bruce D.
A new type of label-free electrochemical immunosensor for the high-sensitivity determination of parathion was developed based on the oriented immobilization of nanobody (VHH9) on a gold nanoparticle-loaded polyvinyl alcohol/citric acid nanofiber membrane-modified electrode. The morphology characterization and assembly process of the modified materials were investigated using scanning electron microscopy (SEM) and electrochemical impedance spectroscopy (EIS). Under the optimum conditions, the label-free electrochemical immunosensor for parathion exhibited a linear range of 0.0015–6400 ng/mL and a low detection limit of 0.48 pg/mL, the signal response of which was 10 times higher than that of the randomly immobilized VHH9. The immunosensor possessed high selectivity, good repeatability and reusability (keeping above 90% of its initial activity after repeating 8 times), and stability (remaining 90% after 9 weeks of storage). Finally, the average recoveries of parathion from food samples were 93.76–105.73% with the coefficient of variation being 2.65–6.85%, showing good correlation with UPLC ( R 2 = 0.9950). Therefore, our nanobody immobilization protocol is simple and effective and proves the potential to be utilized as a promising candidate for sensing platform. Graphical
研究领域:
生物传感
·
环境监测
·
食品检测
✓ 已验证
产品关联
DOI
PubMed
高影响因子
JOURNAL OF HAZARDOUS MATERIALS
·
2025
·
IF 11.3
Yongli Zhang,Zhenyun He,Zhichang Sun,Sihang Zhang,Xing Liu
Ochratoxin A (OTA) poses significant risks to both environment and human health, necessitating the development of rapid and sensitive detection methods. Lateral flow immunosensors (LFIs) typically use monoclonal antibodies and artificial antigens for mycotoxin detection; however, the preparation of these components is time-intensive, laborious, costly, and environmentally harmful. Herein, we developed an innovative genetic engineering-powered colorimetric/fluorescent dual-mode LFI (CM/FL-dLFI) using nanobodies and the mimotope peptide (MP) of OTA. Initially, a heptameric bifunctional fusion (sfGFP-C4bpα-Y4) containing the superfolder green fluorescent protein, the C4-binding protein α-chain, and the MP Y4 was constructed. The fusion was subsequently labeled on gold nanoflowers (AuNFs) to create the AuNFs@sfGFP-C4bpα-Y4 probe, which can be captured by the nanobody heptamer (Nb-C4bpα) and provides colorimetric/fluorescent signals. The optimized CM/FL-dLFI achieved a colorimetric limit of detection (LOD) of 0.01 ng/mL and a fluorescent LOD of 0.05 ng/mL. It exhibited high selectivity for OTA, good recovery rates of 82.85–102.81 % with relative standard deviations not exceeding 14 %, and excellent long-term stability. Moreover, it showed strong correlation with high-performance liquid chromatography in the analysis of 11 real pepper samples. Therefore, this study highlights the promising application potential of nanobodies and MPs in developing an economical and eco-friendly LFI for OTA detection in pepper.
研究领域:
食品科学
·
遗传工程
·
环境健康
✓ 已验证
产品关联
DOI
PubMed
高影响因子
FOOD CHEMISTRY
·
2024
·
IF 8.5
Long Wu,Luyao Jiao,Danni Xue,Yueqing Li,Yu Han,Wei Ouyang,Qi Chen
Microcystin-LR (MC-LR), a potent cyanotoxin in freshwater, poses a risk of severe liver damage and other health issues, making its detection vital. However, the detection capabilities of conventional antibodies are constrained, which limited their use in immunoassays. In this work, we designed a new bifunctional nanobody, named A2.3-SBP (comprised of nanobody and streptavidin binding peptide), capable of binding with MC-LR and streptavidin. Based on A2.3-SBP and Fe 3 O 4 @Au-Pt nanozyme, we introduced an enzyme-free immunosensor that operated in microplate with colorimetric and surface-enhanced Raman scattering (SERS) detection modes. The dual-mode assay showed color changes and SERS intensity directly correlating to MC-LR concentrations with a range from 1.0 to 500 ng/mL and a limit of detection of 0.26 and 0.032 ng/mL, respectively. This strategy eliminated the need for complex enzymatic reactions and realized dual-signal detection of MC-LR in 96 water samples (0.03 μg/kg) within 30 min, suggesting its potential in drinking water detection.
研究领域:
环境科学
·
生物技术
·
分析化学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）