SENSORS AND ACTUATORS B-CHEMICAL
·
2022
·
IF 9.221
Kai Zhang,Meina Yang,Helin Sun,Xueyin Wang,Hao Li,Jinxiang Han,Zhongwen Zhang
Using Peptide nucleic acid (PNA), this work provides a unique approach for controlling CRISPR/Cas13a activity by RNA Inhibitory PNA (RIP). This method reduces the action of CRISPR/Cas13a by inhibiting the binding of crRNA to CRISPR/Cas13a, which was subsequently utilized to detect metalloproteinase-2 (MMP-2) activity. We carefully constructed a PNA sequence that includes a peptide sequence (Gly-Pro-Leu-Gly-Val-Arg-Gly) that is particularly cleaved by MMP-2 in the middle. When MMP-2 is not present in the system, the PNA/crRNA complex reduces crRNA binding to CRISPR/Cas13a, hence inhibiting CRISPR/Cas13a cleavage activity. When MMP-2 is present in the system, it cleaves the peptide sequence (Gly-Pro-Leu-Gly-Val-Arg-Gly) selectively and so prevents PNA from binding to crRNA, enabling crRNA to reach the "free" form. Then, in the presence of an RNA activator, crRNA may bind to CRISPR/Cas13a and stimulate its activity. CRISPR/Cas13a triggered in this manner may precisely cleave the DNA sequence containing modified -UU- bases on the electrode surface . Thus, the ferrocene (Fc) is changed away from the electrode surface at one end of the DNA , enabling for the recovery of the Electrochemiluminescence (ECL) signal from the two-dimensional material transformed on the electrode surface. As a result, MMP-2 activity was detected utilizing this technique of inhibiting crRNA activity with a detection limit of 15.56 fM. Our suggested method of inhibiting CRISPR/Cas13a activity via PNA has a broad variety of potential therapeutic applications in gene editing and the development of CRISPR/Cas13a-based biosensors.
研究领域:
分子生物学
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基因编辑
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生物传感技术
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DOI
高影响因子
BIOSENSORS & BIOELECTRONICS
·
2024
·
IF 10.7
Xiaotian Guan,Jingru Zhao,Zhou Sha,Yujie Liang,Jianghong Huang,Jun Zhang,Shuqing Sun
Exosomes, as novel biomarker for liquid biopsy, exhibit huge important potential value for cancer diagnosis. However, various proteins show different expression levels on exosomal membrane, and the absolute concentration of exosomes in clinical samples is easily influenced by a number of factors. Here, we developed a CRISPR/Cas12a and aptamer-chemiluminescence based analysis (CACBA) for the relative abundance determination of tumor-related protein positive exosomes in plasma for breast cancer diagnosis. The total concentration of exosomes was determined through captured CD63 using a CRISPR/Cas12a-based method with the LoD of 8.97 × 10 3 particles/μl. Meanwhile, EpCAM and MUC1 positive exosomes were quantitatively detected by aptamer-chemiluminescence (ACL) based method with the LoD of 1.45 × 10 2 and 3.73 × 10 2 particles/μl, respectively. It showed that the percentages of EpCAM and MUC1 positive exosomes offered an excellent capability to differentiate breast cancer patients and healthy donors. The high sensitivity, strong specificity, outstanding anti-interference capability, and steady recovery rate of this approach offered higher accuracy and robustness than the commercialized method in clinical trial. In addition with good stability, easy preparation and low cost, this method not only provides a new approach to rapid analysis of exosome proteins, it may be quickly extended to the diagnoses of various cancers.
研究领域:
生物医学工程
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肿瘤学
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分子诊断学
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DOI
PubMed
高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2021
·
IF 7.46
Kai-Ren Zhao,Li Wang,Peng-Fei Liu,Xiao-Min Hang,Hui-Yi Wang,Shu-Ying Ye,Zhi-Jun Liu,Guo-Xi Liang
Herein, we report a “on-off” signal-switchable electrochemiluminescence (ECL) biosensor for simple, sensitive and quantitative detection of two kinds of virus genes, human immunodeficiency virus (HIV) and human papilloma virus (HPV-16) DNAs, by the integration of spherical nucleic acid (SNA) with CRISPR/Cas12a. The concept behind this approach is “on” signal based on a sandwich type assay after ECL luminophores of carbon dots (CDs) labeled SNA incubated with the biosensor in the presence of HIV DNA; and “off” signal upon the biosensor incubation with assistant DNA (HIV DNA sequence), CDs-labeled SNA and Cas12a/crRNA/HPV-16 DNA ternary complex relying on the collaterally cleavage of Cas12a. The cross-interference between HIV and HPV-16 DNAs can be avoided by adding Cas12a effector to the system or not. Using this strategy, HIV and HPV-16 DNAs are successfully quantitatively detection with detection limits of 30 fM and 0.32 pM (S/N = 3), respectively. Importantly, this system does not rely on multiplex signal reporting molecules, thus simplifying the detection system and avoiding the likelihood of cross interferences presented in the assay. Furthermore, multiplex detection of HIV/HPV-16 DNAs in human serum samples are successfully achieved within 2 h without additional nucleic acid amplification step. As demonstrated, the proposed biosensor has the potential for multiplex detection in clinical diagnosis.
研究领域:
生物传感技术
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分子诊断学
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病毒学
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产品关联
DOI
高影响因子
FOOD CHEMISTRY
·
2024
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IF 8.5
Jingrui Yuan,Liu Wang,Fang He,Liang Huang,Kaiyu He,Hongmei Wang,Jing Wang,Xiahong Xu
Salmo salar is one of the most popular salmon species due to its meaty texture and quality protein. Oncorhynchus mykiss , which has a muscle texture similar to that of Salmo salar and is less expensive, is often used as a substitute for Salmo salar . As Salmo salar and Oncorhynchus mykiss belong to the same subfamily of Salmonidae, traditional methods are ineffective in the specific detection of the two. In this study, we combined hue-change with CRISPR/Cas12a lateral flow assay to detect the Salmo salar adulteration. This method detected S. salar genomic DNA at a vLOD of 5 copies, and was able to accurately identify adulterated samples containing 5 % w /w Salmo salar within one hour. In addition, the detection of Salmo salar in processed food products was achieved with the naked-eye at a concentration range of 0 % ∼ 70 % w/w, and the detection accuracy is between 93.3 % ∼ 100 %.
研究领域:
食品科学
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分子生物学
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生物技术
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产品关联
DOI
PubMed
高影响因子
ANALYTICA CHIMICA ACTA
·
2025
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IF 6
Yi Tian,Jiaxuan Chen,Fengzheng Chen,Junru Xu,Lixiang Huang,Liai Peng,Honglei Li,Kai Shi
Background The threat of toxic malathion residues to human health has always been a serious food safety issue. The CRISPR/Cas system represents an innovative detection technology for pesticide residues, but its application to malathion detection has not been reported yet. In addition, the multiple-guide RNA (gRNA) powered-CRISPR/Cas biosensor has the advantages of being fast, sensitive and does not require pre-amplification. However, the reported multiple-gRNA CRISPR/Cas-based biosensors are largely only used for the detection of nucleic acid targets, and there are still certain challenges in detecting non-nucleic acid targets. Results In this work, a multiplex-gRNA-assisted CRISPR/Cas12a-based portable aptasensor (MgCPA) is developed for amplification-free and quantitative detection of malathion using a glucometer. When target malathion is present in the MgCPA strategy, it specifically binds with aptamer and then activates the trans -cleavage activity of the multiplex-gRNA CRISPR/Cas12a. The activated multiple Cas12a/gRNA complexes cut invertase-HP probes on the electrode surface to obtain glucose signals with glucometer assistance. Under optimal conditions, the developed MgCPA strategy achieves satisfactory portable quantitative and sensitive detection of malathion down to 300 fM (S/N = 3) without pre-amplification. Moreover, the satisfactory selectivity, high reproducibility, and good stability of the proposed strategy are also obtained. Due to its excellent and robust shelf life, our developed MgCPA strategy can be practically applied in detecting malathion in orange, apple, cabbage, and spinach samples. Significance Amplification-free, sensitive, portable quantitative and selective detection of malathion in food samples is achieved by employing our developed MgCPA strategy. This strategy not only opens up a new path for the non-nucleic-acid target detection using amplification-free methods based on multiple-gRNA-assisted CRISPR/Cas12a, but also has broad application prospects in ensuring food safety.
研究领域:
生物技术
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食品安全检测
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分子诊断
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产品关联
DOI
PubMed
高影响因子
npj Precision Oncology
·
2025
·Vol. 9
·期 1
·1-21
·
IF 8
Li Xiaoli,Wang Jiashuo,Guo Jiaxing,Zhang Ming
Cisplatin resistance is a major challenge in laryngeal cancer treatment. Tumor necrosis factor-alpha-induced protein 2 (TNFAIP2) has been implicated in chemoresistance, though its role in laryngeal cancer remains unclear. We identified TNFAIP2 as a key gene associated with cisplatin resistance and developed a second near-infrared (NIR-II) light-responsive CRISPR-Cas9 nanosystem (APC@RBCs) to target it. In cisplatin-resistant Tu177/CDDP cells, light-activated knockout of TNFAIP2 significantly increased cisplatin sensitivity (IC50 reduced from 12.55 to 4.37 µg/mL). Mechanistic studies revealed that TNFAIP2 modulates cisplatin resistance via the NRF2 pathway, affecting oxidative stress response and epithelial-mesenchymal transition. TNFAIP2 deletion also suppressed cancer cell migration and invasion. Our results establish TNFAIP2 as a critical mediator of chemoresistance and demonstrate the potential of APC@RBCs as a precise, non-invasive gene therapy platform. This strategy offers a promising approach to overcoming cisplatin resistance and improving treatment outcomes in laryngeal cancer patients.
研究领域:
肿瘤学
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分子生物学
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药理学
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纳米技术
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基因工程
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产品关联
DOI
PubMed
高影响因子
BIOSENSORS & BIOELECTRONICS
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2022
·
IF 12.545
Hai-Yan Jia,Hong-Li Zhao,Ting Wang,Pin-Ru Chen,Bin-Cheng Yin,Bang-Ce Ye
MicroRNAs (miRNAs) play an essential role in cancer diagnosis and prognosis. Developing a new method for sensitive detection of miRNA is constantly in demand. CRISPR/Cas12a system can nonspecifically cleave single-stranded DNA after specific recognition of target DNA, showing tremendous potential in molecular diagnostics. However, CRISPR-based detection methods require synthesizing different crRNAs for detecting different targets, which limit their widespread application. Herein, we design a versatile and sensitive miRNA detection platform based on CRISPR/Cas12a system combined with a hybridization chain reaction (HCR) circuit. In this design, the HCR circuit as the signal transducer converts each miRNA into multiple DNA duplexes, which act as the activators to activate the trans -cleavage activity of Cas12a for further signal amplification . More importantly, this platform can sensitively detect different miRNAs without changing the spacer sequence of crRNA due to the fixed activators formed by HCR. In addition, the consistency between the proposed platform and RT-qPCR in miRNA detection extracted from different cell lines validated its practicability, demonstrating the potential in clinical diagnosis of cancers and monitoring therapy.
研究领域:
分子生物学
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生物技术
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癌症研究
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诊断技术
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产品关联
DOI
PubMed
高影响因子
Advanced Science
·
2025
·
IF 14.1
Yuwei Li,Yuanyi Zhang,Chenchen Li,Guoping Chen,Pir Muhammad,Yonghong Yao,Lifang Gao,Zhigang Liu,Yanli Wang
Cancer immunotherapy that activates the stimulator of interferon genes (STING) signaling pathway to resist tumors has recently attracted considerable attention. However, STING activation can induce opposing interferon functions that contribute to T-cell exhaustion via programmed death-ligand 1 (PD-L1). In particular, effectively using the immune system to combat tumors remains a substantial challenge due to tumor immunosuppressive factors such as SMARCAL1. Here, a glucose-responsive CRISPR nanovaccine is developed for enhancing STING signaling while inhibiting interferon-mediated immunosuppressive feedback. The formulation encapsulates a bimetallic zeolitic imidazolate framework with glucose oxidase (GOx) and CRISPR-mediated SMARCAL1 gene-editing plasmids. The dual enzyme-driven cascade reactions of peroxidase and GOx generate reactive oxygen species (ROS) and gluconic acid, which release and activate the genome-editing system. The silencing of SMARCAL1 enhances STING activity and inhibits PD-L1 expression, resulting in the termination of PD-L1-mediated opposing functions of interferon. Zinc ions and double-stranded DNA formed via ROS further activate the STING pathway, effectively inducing dendritic cell maturation and immune system activation. This is a critical report of in situ CRISPR nanovaccination driven by dual enzymes. The work highlights the potential of glucose-responsive CRISPR nanovaccination in bolstering antitumor immunity and extends the implementation of gene editing in cancer immunotherapy.
研究领域:
免疫治疗
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肿瘤学
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基因工程
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纳米医学
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生物化学
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分子生物学
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产品关联
DOI
PubMed
高影响因子
ANALYST
·
2022
·Vol. 147
·期 24
·5687-5693
·
IF 5.227
Yuyuan Zhang,Jing Lei,Wei Liu,Yan Jin,Baoxin Li
Various CRISPR/Cas12a-based biosensing systems have been developed in the past few years, and most of these systems used Taqman probes to report signals through fluorescence resonance energy transfer (FRET). In this study, we explored chemiluminescence resonance energy transfer (CRET) as the readout mode for CRISPR/Cas12a-based biosensing. The chemiluminescence (CL) reaction of bis(2,4,6-trichlorophenyl) oxalate (TCPO) and H2O2 was used to excite the fluorophore dye of the Taqman probe. Different from FRET, CRET does not need external excitation light, which can effectively avoid autofluorescence and photobleaching. The detection limit of this CRET readout mode was estimated to be 10 pM for target DNA, which was about 8 times lower than that of the widely used FRET readout mode. These results suggest that CRET can serve as a rapid, sensitive and simple readout mode of CRISPR/Cas12-based biosensing, and can further enrich the toolbox of CRISPR/Cas12-based biosensing.
研究领域:
分子生物学
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生物化学
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诊断学
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产品关联
DOI
PubMed
高影响因子
Theranostics
·
2025
·Vol. 15
·期 7
·2870
·
IF 13.3
Xuena Zhu,Shanshan Wang,Yuanyuan Xue,Xiaoyan Wang,Shaoqi Hu,Tingbo Liang,Wenjun Liu
Significant efforts and resources have been dedicated to developing CRISPR-Cas based point-of-care testing (POCT) and self-diagnosis methods for nucleic acid pathogens, aiming to complement the gold standard quantitative PCR tests, particularly in settings where centralized facilities, trained personnel, or resource-intensive equipment are unavailable. However, the reliance on stationary, high-cost readout machinery hinders their full deployment at the point of care. We aimed to develop a solid-phase invertase-labeled reporter (ILR) system that enables convenient readout of CRISPR-Cas reactions, facilitate HPV detection in a POCT-compatible manner. Methods: Through multiple chemical couplings, invertase is immobilized onto magnetic microbeads via a nucleic acid linker that responds to target nucleic acid-induced CRISPR-Cas activation. This activation releases active invertase, which then converts sucrose to glucose in proportion to the target's abundance. Enzymatic signal amplification by Cas12a/Cas13a and invertase compensates for the moderate sensitivity of personal glucose meters (PGMs). Results: When applied to human papillomavirus detection, the HPV18-targeted LAMP-Cas12a/ILR/PGM system can detect as few as 7 HPV18-positive HeLa cells out of 7,000, achieving 95.8% sensitivity and 100% specificity in cervical cell samples. Furthermore, minimal reagent adjustments allow for the rapid establishment of HPV16 and HPV52-targeted LAMP-Cas12a/ILR/PGM systems, offering satisfactory sensitivity, specificity, and cross-species detection. Conclusion: These findings demonstrate a highly efficient testing platform for a range of nucleic acid pathogens, suitable for both point-of-care and household use.
研究领域:
分子生物学
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医学诊断
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生物化学
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产品关联
DOI
PubMed
高影响因子
CANCER RESEARCH
·
2025
·
IF 16.6
Shang Yan,Li Shuaihu,Xu Tian,Sun Yihua,Li Fei
The CRISPR/Cas9 lentiviral system has emerged as a powerful tool for gene knockout in cancer immunology research. However, the persistent expression of exogenous elements, such as Cas9 and resistance markers, often triggers excessive tumor immune rejection. This can lead to prolonged experimental timelines, increased data variability, biased outcomes, and even experimental failures. To address this challenge, several strategies have been investigated that offer partial solutions, but they have failed so far to comprehensively resolve the issue. In response, we developed the VL-AdCre system, a strategy that enables efficient excision of exogenous expression elements following gene knockout. The VL-AdCre system effectively reduced tumor immune rejection in allograft models, streamlined experimental workflows, and improved the reliability of research outcomes. Overall, this optimized CRISPR/Cas9 lentiviral gene knockout system offers a robust and practical solution for studying gene functions in vivo and advancing immunotherapeutic strategies.
研究领域:
免疫学
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肿瘤学
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分子生物学
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基因编辑
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转化医学
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产品关联
DOI
PubMed
高影响因子
TALANTA
·
2024
·
IF 5.6
Yu Tao,Haixia Wang,Enguo Ju,Yeh-Hsing Lao,Yu Zhang,Mingqiang Li
Diseases caused by viruses, such as monkeypox virus (MPXV) and human papillomavirus (HPV), pose serious threats to human health and safety. Although numerous strategies have been constructed for detecting MPXV and HPV DNA, most methods require either laborious procedures or complicated instruments involving skilled professionals. In this research, a CRISPR-Cas12a-mediated colorimetric detection platform for MPXV and HPV sensing was constructed for the first time by applying probe DNA to reprogram the catalytic properties of molybdenum disulfide quantum dots (MoS 2 QDs). In the presence of MPXV or HPV targets, the CRISPR-Cas12a trans -cleavage activity is effectively motivated to decompose the probe DNA, leading to the suppression of enzymatic activity DNA enhancer adsorbed on MoS 2 QDs, resulting in greatly decreased catalytic behaviors. The MoS 2 QDs-DNA nanohybrids displayed prominent specificity and sensitivity, with detection limits at subpicomolar levels, as well as excellent stability and accuracy for determining MPXV and HPV DNA in human sera biosamples. Furthermore, the proposed colorimetric biosensing approach can be ensembled with a smartphone platform, allowing visible analysis of DNA targets. Taken together, this colorimetric strategy offers a novel diagnosis method for MPXV and HPV DNA detection, particularly favorable for highly endemic developing countries with restricted medical and instrumental support.
研究领域:
分子生物学
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病毒学
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医学诊断
✓ 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）