Biosensors-Basel
·
2024
·Vol. 14
·期 3
·135
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IF 4.9
Hui Chen,Zhiyuan Zhuang,Naihan Xu,Ying Feng,Kaixin Fang,Chunyan Tan,Ying Tan
The ongoing COVID-19 pandemic, driven by persistent SARS-CoV-2 transmission, threatens human health worldwide, underscoring the urgent need for an efficient, low-cost, rapid SARS-CoV-2 detection method. Herein, we developed a point-of-care SARS-CoV-2 detection method incorporating recombinase polymerase amplification (RPA) and DNA–protein crosslinking chemiluminescence (DPCL) (RPADPCL). RPADPCL involves the crosslinking of biotinylated double-stranded RPA DNA products with horseradish peroxidase (HRP)-labeled streptavidin (SA-HRP). Modified products are captured using SA-labeled magnetic beads, and then analyzed using a chemiluminescence detector and smartphone after the addition of a chemiluminescent substrate. Under optimal conditions, the RPADPCL limit of detection (LOD) was observed to be 6 copies (within the linear detection range of 1–300 copies) for a plasmid containing the SARS-CoV-2 N gene and 15 copies (within the linear range of 10–500 copies) for in vitro transcribed (IVT) SARS-CoV-2 RNA. The proposed method is convenient, specific, visually intuitive, easy to use, and does not require external excitation. The effective RPADPCL detection of SARS-CoV-2 in complex matrix systems was verified by testing simulated clinical samples containing 10% human saliva or a virus transfer medium (VTM) spiked with a plasmid containing a SARS-CoV-2 N gene sequence or SARS-CoV-2 IVT RNA. Consequently, this method has great potential for detecting targets in clinical samples.
研究领域:
分子诊断
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病毒学
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生物医学工程
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分析化学
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产品关联
DOI
PubMed
高影响因子
CHEMICAL ENGINEERING JOURNAL
·
2026
·
IF 13.2
Lijuan Yin,Liangyuan Wang,Lu Yin,Lingzhi Yang,Baoxue Gao,Shuli Man,Fufeng Liu,Long Ma
Norovirus (NoV) represents a leading cause of foodborne illness worldwide and is the predominant agent of non-bacterial acute gastroenteritis. The development of rapid, highly sensitive, specific, and point-of-care detection methods is critical for the protection of public health and food safety. In this study, we developed a lyophilized paper-based microfluidic analytical device (μPAD) that integrates reverse transcription recombinase polymerase amplification (RT-RPA) with CRISPR/Cas12a technology. By incorporating a smartphone-assisted portable fluorescence detection device, the platform achieves a high level of integration, simplicity, and intelligence. It demonstrates a detection limit for NoV as low as 1 copy/μL within 45 min, with no cross-reactivity against other common microorganisms or the human genome. The fluorescence intensity on the chips exhibits a strong linear relationship with NoV RNA concentration, enabling accurate quantification. Moreover, the lyophilized RPA-CRISPR paper chips retain stability for up to 8 weeks at room temperature, underscoring their suitability for point-of-care applications. When applied to real food and environmental samples for NoV detection, the platform showed 100% concordance with RT-qPCR and maintained a detection sensitivity of 1 copy/μL. This work provides a robust approach for rapid, sensitive, and on-site detection of norovirus, and the platform architecture holds promise for broader application in detecting other microorganisms.
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DOI
高影响因子
Food Bioscience
·
2026
·
IF 6.2
Jiali Wang,Yuting Su,Yaru Chen,Lei Tang,Dongmei Li,Wenchao Du,Yumin Shen,Fangfang Zhou,Hong Duan,Baoguo Sun,Gang Chen,Lingyan Zheng
Burkholderia gladioli pv . cocovenenans (BGC) is an emerging foodborne pathogen capable of producing the deadly toxins bongkrekic acid and toxoflavin, posing significant risks to public health. Currently, rapid screening for this bacterium is severely challenged by the inherently complex nature of various food matrices and the low throughput of routine nucleic acid extraction. Here we report a pipette-tip-compatible annular magnetic separator (AMS) that efficiently operates with magnetic silica beads in 96-well plate format. This innovative device enables parallel DNA extraction of 96 samples within merely 13.5 min without heating or centrifugation, thus streamlining the pretreatment process. This platform integrates the high-throughput extraction step with real-time fluorescent recombinase polymerase amplification (RT-RPA) specifically targeting the Bon M gene for the rapid detection of BGC in food products. Under optimized conditions, the integrated AMS-RT-RPA platform provides extraction-to-detection diagnostic results in 33.5 min (comprising 13.5 min for DNA extraction and approximately 20 min for RT-RPA amplification), and achieves a limit of detection as low as 190 CFU/mL. Furthermore, the spiked samples of black fungus and rice flour were also tested by the detection platform, and the recoveries ranged from 89.7% to 103.2%, with the coefficient of variation less than 11.6%. Collectively, this work demonstrates the practical feasibility of a scalable pretreatment-amplification workflow, highlighting its potential for high-throughput on-site screening during foodborne emergency investigations and routine food safety monitoring.
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DOI
高影响因子
MICROCHEMICAL JOURNAL
·
2025
·
IF 5.1
Ying Cheng,Jiaxuan Chen,Jiuhua Song,Jiali Chang,Yang Li,Qiao Zhang,Yifan Feng,Kai Shi,Tao Jiang
Detection of MPXV DNA in urban wastewater is crucial for early warning, rapid response, and effective outbreak control, offering a non-invasive approach to strengthen epidemic preparedness and public health defense. Based on heparin sodium-assisted one-tube recombinase polymerase amplification (RPA)/split gRNA with Cas12a (SCas12a) and phosphorothioate-modified hairpin G-quadruplex (psHG4), we have developed a colorimetric biosensing platform to achieve sensitive and on-site detection of MPXV DNA in urban wastewater (called hoRPA/SCas12a). The MPXV DNA initiates a one-tube RPA/SCas12a reaction facilitated by heparin sodium, subsequently inducing the DNase activity of SCas12a. The loop DNA of psHG4 is cleaved by active SCas12a to release the pre-blocked psG4 sequence, enabling the formation of a psG4/hemin complex with K + and hemin. The complex is capable of initiating the oxidation of 3,3′, 5,5′-tetramethylbenzidine (TMB) to generate the colorimetric substrate TMBox. The portable smartphone was employed to record the RGB value changes of the sample. The hoRPA/SCas12a platform displays a linear range of 10 aM to 100 fM, with a detection limit of 3 aM. Moreover, this platform can selectively and accurately detect MPXV DNA in urban wastewater. The hoRPA/SCas12a platform is vital for early warning, effective public health response, and strengthening wastewater-based epidemiology in emerging infectious disease control.
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产品关联
DOI
高影响因子
CHEMICAL ENGINEERING JOURNAL
·
2024
·
IF 13.3
Haoyu Wang,Shiting Chen,Jiqing Liu,Wanyue Liu,Wei Chen,Lin Huang,Chao Yan
In recent years, RPA-CRISPR/Cas12a-based electrochemical biosensors have emerged as highly effective tools for nucleic acid detection. However, the random trans -cleavage characteristics of Cas12a on single-stranded nucleic acid sequences can result in the presence of uncertain nucleic acid residues on the electrode surface post-reaction. This situation complicates subsequent sensitization strategies. In this study, we devised a universal sensitization protocol mediated by terminal deoxynucleotidyl transferase (TdT) , a template-independent DNA polymerase and G-rich spherical nucleic acids (G-rich SNAs) with superior performance. Initially, Cas12a was activated by the specific RPA amplification product and cleaved the closed single stranded DNA (C-ssDNA) on the electrode surface. Then, the TdT enzyme extended residual single stranded DNA (R-ssDNA) on the electrode surface, forming G-Quadruplex structures. Subsequently, G-rich SNAs were immobilized on the electrode surface via π-π stacking interactions with the G-quadruplexes. Consequently, a substantial accumulation of hemin occurred within the G-quadruplex cluster structures, generating significant electrochemical signals. We applied this method to the trace detection of Staphylococcus aureus ( S. aureus ) and achieved a good linear calibration curve with concentrations ranging from 0 CFU/mL to 10 4 CFU/mL. Notably, the limit of detection (LOD) was as low as 1.29 CFU/mL, surpassing that of most similar sensors. These results demonstrate that our proposed universal sensitization method effectively addresses the limitations associated with the random trans -cleavage characteristics of Cas12a.
研究领域:
生物传感
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纳米技术
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分子生物学
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电化学
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产品关联
DOI
高影响因子
PROCESS BIOCHEMISTRY
·
2023
·
IF 4.4
Xiaoxuan Cai,Miaoen Huang,Xixiang Huang,Huan Liu,Tianji Wang,Li Li,Weiguang Yang,Hui Luo,Yingnian Lu
The organism Sipunculus nudus L is a good resource for marine peptides because of its rich protein. However, the disadvantage of biological peptides is their short metabolic half-life, therefore, structural modification of peptides is necessary. In this research, the enzymatic hydrolysis of Sipunculus nudus L was performed using trypsin and then peptides were isolated and purified from the hydrolysates, four novel tripeptides were identified by LC-MS-MS as Ser-Arg-Pro (SRP, m/z = 358.39), Pro-Arg-Pro (PRP, m/z = 368.43), Arg-Pro-Ala (RPA, m/z = 342.39), and Lue-Pro-Lys (LPK, m/z = 356.46), their ACE inhibitory activity was investigated with IC 50 value as following 0.046, 0.42, 0.97, and 6.54 mM, respectively. Because of its highest ACE inhibitory activity, SRP was further selected to modify the chemical structure for a sustained release agent DSPE-PEG-SRP. Compared with 7 h of free SRP, the release of DSPE-PEG-SRP in vitro was approximately 120 h. The ACE inhibition rate of SRP released from the synthetic product and the raw material SRP was almost the same, molecular docking demonstrated that ACE inhibitory activity was attributed to the formation of hydrogen bonds between SRP and ACE active sites . In conclusion, DSPE-PEG-SRP, a novel sustained release agent, has great potential and is expected to be further applied to treat hypertension.
研究领域:
海洋生物学
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药学
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生物化学
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DOI
高影响因子
BIOSENSORS & BIOELECTRONICS
·
2021
·
IF 10.618
Zhiying Wang,Yang Wang,Long Lin,Tao Wu,Zhenzhen Zhao,Binwu Ying,Lingqian Chang
Tuberculosis (TB) remains the high-risk infectious pathogen that caused global pandemic and high mortality, particularly in the areas lack in health resources. Clinical TB screening and diagnosis are so far mainly conducted on limited types of commercial platforms, which are expensive and require skilled personnel. In this work, we introduced a low-cost and portable finger-driven microfluidic chip (named Fd-MC) based on recombinase polymerase amplification (RPA) for rapid on-site detection of TB. After injection of the pre-treated sample solution, the pre-packaged buffer was driven by the pressure generated by the finger-actuated operation to accomplish sequential processes of diagnosis in a fully isolated microchannel . An in-situ fluorescence strategy based on FAM-probe was implemented for on-chip results read-out though a hand-held UV lamp. Hence, the Fd-MC proved unique advantageous for avoiding the risk of infection or environmental contamination. In addition, the Fd-MC was designed for multiplexed detection, which is able to not only identify TB/non-TB infection, but also differentiate between human Mycobacterium tuberculosis and Mycobacterium bovis . The platform was verified in 37 clinical samples, statistically with 100% specificity and 95.2% sensitivity as compared to commercial real-time RPA. Overall, the proposed platform eliminates the need on external pumps and skilled personnel, holding promise to POC testing in the resource-limited area.
研究领域:
生物医学工程
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微流控技术
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分子诊断
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传染病
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即时检测
·
全球健康
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产品关联
DOI
PubMed
高影响因子
Cell Reports
·
2024
·
IF 7.5
Sijia Ren,Feng Bai,Viviane Schnell,Clara Stanko,Muriel Ritsch,Tino Schenk,Emanuel Barth,Manja Marz,Bin Wang,Xin-Hai Pei,Holger Bierhoff
Extensive remodeling of the female mammary epithelium during development and pregnancy has been linked to cancer susceptibility. The faithful response of mammary epithelial cells (MECs) to hormone signaling is key to avoiding breast cancer development. Here, we show that lactogenic differentiation of murine MECs requires silencing of genes encoding ribosomal RNA (rRNA) by the antisense transcript PAPAS. Accordingly, knockdown of PAPAS derepresses rRNA genes, attenuates the response to lactogenic hormones, and induces malignant transformation. Restoring PAPAS levels in breast cancer cells reduces tumorigenicity and lung invasion and activates many interferon-regulated genes previously linked to metastasis suppression. Mechanistically, PAPAS transcription depends on R-loop formation at the 3' end of rRNA genes, which is repressed by RNase H1 and replication protein A (RPA) overexpression in breast cancer cells. Depletion of PAPAS and upregulation of RNase H1 and RPA in human breast cancer underpin the clinical relevance of our findings.
研究领域:
癌症生物学
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分子生物学
·
细胞生物学
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DOI
PubMed
高影响因子
BIOSENSORS & BIOELECTRONICS
·
2022
·
IF 12.545
Xiangqin Meng,Sijia Zou,Dandan Li,Jiuyang He,Ling Fang,Haojue Wang,Xiyun Yan,Demin Duan,Lizeng Gao
The coronavirus disease 2019 (COVID-19) pandemic has created a huge demand for sensitive and rapid detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The current gold standard for SARS-CoV-2 detection is reverse transcription-polymerase chain reaction (RT-PCR)-based nucleic acid amplification. However, RT-PCR is time consuming and requires specialists and large instruments that are unattainable for point-of-care testing (POCT). To develop POCT for SARS-CoV-2, we combined recombinase polymerase amplification (RPA) and FeS 2 nanozyme strips to achieve facile nucleic acid amplification and subsequent colorimetric signal enhancement based on the high peroxidase-like activity of the FeS 2 nanozymes. This method showed a nucleic acid limit of detection (LOD) for SARS-CoV-2 of 200 copies/mL, close to that of RT-PCR. The unique catalytic properties of the FeS 2 nanozymes enabled the nanozyme-strip to amplify colorimetric signals via the nontoxic 3,3′,5,5′-tetramethylbenzidine (TMB) substrate. Importantly, the detection of clinical samples of human papilloma virus type 16 (HPV-16) showed 100% agreement with previous RT-PCR results, highlighting the versatility and reliability of this method. Our findings suggest that nanozyme-based nucleic acid detection has great potential in the development of POCT diagnosis for COVID-19 and other viral infections.
研究领域:
分子生物学
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病毒学
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即时诊断
·
纳米技术
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DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）