TRENDS IN BIOTECHNOLOGY
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2025
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IF 14.9
Tian Yu,Siyi Zou,Yingxi Long,Yourong Ou,Shixian Liu,Tongjia Kang,Libing Song,Congcong Sun,Guozhen Liu
Developing a generic sensitive platform for detecting diverse biomarkers is essential for a comprehensive understanding of disease states, guiding precision medicine. Herein, we introduce a versatile platform based on glass fiber interfaced CRISPR/Cas with a universal reagent setting (g-CURS), which used a fixed pair of CRISPR RNA (crRNA) and a single-stranded DNA (ssDNA) activator to enable detection of multiple nucleic acids or proteins with ultrahigh sensitivity. The fixed ssDNA activator was labeled on multiple specific ligation products or detection antibodies conjugated on glass fiber to initiate CRISPR/Cas12a-assisted rapid and exponential cascade amplification through circular reporters (CRs), generating fluorescence signals readable by a portable detector. g-CURS was able to detect viral nucleic acids with attomolar sensitivity within 30 min and multiple low-abundance proteins in extracellular vesicles of Parkinson’s disease (PD) serum with subpicomolar sensitivity within 80 min. g-CURS simplifies CRISPR/Cas biosensing using a standard reagent setting, holding promise for biomarker discovery free from bulky instruments.
研究领域:
分子生物学
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医学诊断
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生物传感技术
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产品关联
DOI
PubMed
高影响因子
ANALYTICAL CHEMISTRY
·
2023
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IF 7.4
Liping Xia,Juxin Yin,Jianjian Zhuang,Weihong Yin,Zheyu Zou,Ying Mu
Rapid and sensitive pathogen detection methods are critical for disease diagnosis and treatment. RPA-CRISPR/Cas12 systems have displayed remarkable potential in pathogen detection. A self-priming digital PCR chip is a powerful and attractive tool for nucleic detection. However, the application of the RPA-CRISPR/Cas12 system to the self-priming chip still has great challenges due to the problems of protein adsorption and two-step detection mode of RPA-CRISPR/Cas12. In this study, an adsorption-free self-priming digital chip was developed and a direct digital dual-crRNAs (3D) assay was established based on the chip for ultrasensitive detection of pathogens. This 3D assay combined the advantages of rapid amplification of RPA, specific cleavage of Cas12a, accurate quantification of digital PCR, and point-of-care testing (POCT) of microfluidics, enabling accurate and reliable digital absolute quantification of Salmonella in POCT. Our method can provide a good linear relationship of Salmonella detection in the range from 2.58 × 101 to 2.58 × 104 cells/mL with a limit of detection ∼0.2 cells/mL within 30 min in a digital chip by targeting the invA gene of Salmonella. Moreover, the assay could directly detect Salmonella in milk without nucleic acid extraction. Therefore, the 3D assay has the significant potential to provide accurate and rapid pathogen detection in POCT. This study provides a powerful nucleic detection platform and facilitates the application of CRISPR/Cas-assisted detection and microfluidic chips.
研究领域:
分子生物学
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生物技术
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微生物学
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分析化学
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即时诊断
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产品关联
DOI
PubMed
高影响因子
Biomolecules
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2025
·Vol. 15
·期 3
·315
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IF 4.8
Dongxia Pan,Mukaddas Mijit,Hui Wang,Chaoqun Sun,Bantan Pingcuo,Zhixue Yu,Benhai Xiong,Xiangfang Tang
The A-to-G mutation (FecB) in theBMPR1Bgene is strongly linked to fertility in sheep, significantly increasing ovulation rates and litter sizes compared to wild-type populations. The rapid and reliable screening of theFecBgene is therefore critical for advancing sheep breeding programs. This study aimed to develop a fast and accurate method for detecting theFecBmutation and genotyping the gene to enhance sheep reproduction and productivity. To achieve this, we integrated the CRISPR-Cas12a system with an optimized amplification refractory mutation system (ARMS). A similar DNA origami technique-based fluorescence reporter nanotree structure was synthesized using gold nanomagnetic beads as carriers to amplify the fluorescence signal further. The resulting biosensing platform, termed CRISPR-ARMS, demonstrated excellent sensitivity for detectingFecBmutations, with a detection limit as low as 0.02 pmol. Therefore, this innovative approach shows great promise for single-base mutation detection and represents a pioneering tool for high-yield genetic screening.
研究领域:
分子生物学
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遗传学
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动物科学
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DOI
PubMed
高影响因子
Cell Reports Medicine
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2025
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IF 10.6
Jianhai Chen,Jianwei Tan,Nannan Wang,Hui Li,Wenxiang Cheng,Jian Li,Benguo Wang,Adam C. Sedgwick,Zhitong Chen,Guojun Chen,Peng Zhang,Wei Zheng,Chengbo Liu,Jingqin Chen
Rheumatoid arthritis (RA) is the most prevalent inflammatory arthritis with unknown etiology, characterized by synovial inflammation and articular bone erosion. Studies have highlighted that inhibiting macrophage-induced osteoclastogenesis holds promise in mitigating bone destruction. However, specifically halting this pathological cascade remains a challenge for the management of RA. Here, initially, we identify that Ras homolog gene family member A ( RhoA ) is a pivotal target in inducing osteoclastogenesis of macrophages. Subsequently, we develop a strategy termed specific macrophages RhoA targeting (SMART), in which phosphatidylserine (PS)-enriched macrophage membranes are engineered to deliver macrophage-specific promoter-containing CRISPR-Cas9 plasmids (SMART-Cas9), enabling targeted editing of RhoA in RA joint macrophages. Multiscale imaging techniques confirm the highly specific targeted effect of SMART-Cas9 on the macrophages of inflamed joints. SMART-Cas9 successfully reduces osteoclastogenesis by macrophages, thus mitigating bone erosion by modulating cytoskeletal dynamics and immune balance in inflammatory arthritis, representing a therapeutic avenue for RA and other inflammatory bone diseases.
研究领域:
免疫学
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分子生物学
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风湿病学
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产品关联
DOI
PubMed
高影响因子
TALANTA
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2025
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IF 6.1
Xiuyu Chen,Dan Zhao,Changmin Yu,Jifu Wei,Guoren Zhou
The detection of specific protein biomarkers holds significant potential for the early diagnosis of colorectal cancer (CRC). However, the accurate quantification of low-abundance proteins in serum presents a major challenge due to factors such as limited sensitivity and the complexity of the required methodologies. In this work, we established a universal CRISPR/Cas biosensing platform by integrating novel photosensitive nanoprobes (DA/PL@Cu NPs) and CRISPR/Cas12 system (DPC-Cas) for the highly sensitive, specific and user-friendly detection of angiopoietin-like protein 2 (ANGPTL2). The DA/PL@Cu NPs serve as a critical component in the transduction of protein recognition information into nucleic acid amplification events to produce Cas12a activators. The DPC-Cas biosensor integrates DA/PL@Cu NPs-assisted amplification with Cas12a self-amplification, enabling ultrasensitive detection of ANGPTL2 at concentrations as low as 20.00 pg/mL. The proposed DPC-Cas biosensor successfully detected ANGPTL2 in serum, demonstrating significant potential for the early diagnosis of CRC.
研究领域:
生物医学
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分子诊断
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纳米技术
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产品关联
DOI
PubMed
高影响因子
Nano Today
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2021
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IF 20.722
Dagan Zhang,Lijun Cai,Xiaowei Wei,Yuetong Wang,Luoran Shang,Lingyun Sun,Yuanjin Zhao
Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR associated nucleases (CRISPR/Cas) platforms have a demonstrated value in nucleic acid detection, while their multiplex screening capability is still challenging. Herein, we present a novel CRISPR/Cas9 platform based on bioinspired photonic crystal (PhC) barcodes for multiplex and sensitive nucleic acid detection. The bioinspired PhC barcodes not only exhibit distinctive structural color as encoding elements, but also possess abundant functional surface groups for probe immobilization with the presence of polydopamine (PDA) coating. The CRISPR/Cas9 system recognizes and cleaves target DNA and with the help of Klenow fragment to produce ssDNA, which was subsequently captured by the PhC barcodes and the detection signal was amplified by hybridization chain reaction (HCR). Based on this platform, we demonstrated that Human Papilloma Virus (HPV) nucleic acids could be detected with a highly sensitive limit of 0.025 pM and be multiplexed assayed with high accuracy and specificity. Thus, we believe that this platform paves an avenue for multiplexed biomarkers quantification in clinical disease diagnostics.
研究领域:
分子生物学
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诊断学
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生物医学工程
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产品关联
DOI
高影响因子
Research
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2022
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IF 11.036
Liangfen Cheng,Fuhan Yang,Longfei Tang,Lelin Qian,Xu Chen,Feng Guan,Juan Zhang,Genxi Li
Evaluating tumor development is of great importance for clinic treatment and therapy. It has been known that the amounts of sialic acids on tumor cell membrane surface are closely associated with the degree of cancerization of the cell. So, in this work, cellular interface supported CRISPR/Cas trans-cleavage has been explored for electrochemical simultaneous detection of two types of sialic acids, i.e., N-glycolylneuraminic acid (Neu5Gc) and N-acetylneuraminic acid (Neu5Ac). Specifically, PbS quantum dot-labeled DNA modified by Neu5Gc antibody is prepared to specifically recognize Neu5Gc on the cell surface, followed by the binding of Neu5Ac through our fabricated CdS quantum dot-labeled DNA modified by Sambucus nigra agglutinin. Subsequently, the activated Cas12a indiscriminately cleaves DNA, resulting in the release of PbS and CdS quantum dots, both of which can be simultaneously detected by anodic stripping voltammetry. Consequently, Neu5Gc and Neu5Ac on cell surface can be quantitatively analyzed with the lowest detection limits of 1.12 cells/mL and 1.25 cells/mL, respectively. Therefore, a ratiometric electrochemical method can be constructed for kinetic study of the expression and hydrolysis of Neu5Gc and Neu5Ac on cell surface, which can be further used as a tool to identify bladder cancer cells at different development stages. Our method to evaluate tumor development is simple and easy to be operated, so it can be potentially applied for the detection of tumor occurrence and development in the future.
研究领域:
生物医学工程
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分子生物学
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肿瘤学
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DOI
高影响因子
ANALYST
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2025
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IF 3.3
Shaoying He,Wei Lin,Xin Liu,Fei Li,Hong Liang,Huo Xu,Chunhua Lu,Chao Xing
Lead pollution presents a significant threat to ecological systems and human health, underscoring the urgent need for highly sensitive detection methods. Herein, we introduce a novel DNA concatemer-encoded CRISPR/Cas12a fluorescence sensor (MDD-Cas12a) for sensitive detection of Pb2+ based on DNAzymes. To accomplish this, we designed a substrate strand containing a long DNA concatemer encoding multiple protospacer adjacent motifs (PAMs) and protospacer sequences for activation of the CRISPR/Cas12a system. The DNA concatemer was subsequently anchored to the surface of magnetic beads (MBs) to fabricate a MBs-DNA concatemer nanoprobe. In the presence of Pb2+, the DNAzyme structure catalyzes the cleavage of the substrate strand, leading to the release of DNA concatemers. Following magnetic separation, the released DNA concatemers significantly activate the non-specific trans-cleavage activity of the Cas12a/crRNA complex. The fluorescence reporter DNA is then completely cleaved by the activated Cas12a/crRNA complex, and the Pb2+ concentration in the sample can be quantified by measuring the fluorescence signal. By harnessing the specific recognition capability of DNAzymes for Pb2+, the programmability of DNA concatemers, and the self-amplification features of the CRISPR/Cas12a system, the MDD-Cas12a platform demonstrates high sensitivity and specificity for detecting Pb2+ in milk and lake water samples.
研究领域:
生物化学
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分子生物学
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环境科学
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产品关联
DOI
PubMed
高影响因子
Theranostics
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2022
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IF 11.6
Bohan Yin,Qin Zhang,Xinyue Xia,Chuanqi Li,Willis Kwun Hei Ho,Jiaxiang Yan,Yingying Huang,Honglian Wu,Pui Wang,Changqing Yi,Jianhua Hao,Jianfang Wang,Honglin Chen,Siu Hong Dexter Wong,Mo Yang
Background: CRISPR-Cas12a has been integrated with nanomaterial-based optical techniques, such as surface-enhanced Raman scattering (SERS), to formulate a powerful amplification-free nucleic acid detection system. However, nanomaterials impose steric hindrance to limit the accessibility of CRISPR-Cas12a to the narrow gaps (SERS hot spots) among nanoparticles (NPs) for producing a significant change in signals after nucleic acid detection. Methods: To overcome this restriction, we specifically design chimeric DNA/RNA hairpins (displacers) that can be destabilized by activated CRISPR-Cas12a in the presence of target DNA, liberating excessive RNA that can disintegrate a core-satellite nanocluster via toehold-mediated strand displacement for orchestrating a promising “on-off” nucleic acid biosensor. The core-satellite nanocluster comprises a large gold nanoparticle (AuNP) core surrounded by small AuNPs with Raman tags via DNA hybridization as an ultrabright Raman reporter, and its disassembly leads to a drastic decrease of SERS intensity as signal readouts. We further introduce a magnetic core to the large AuNPs that can facilitate their separation from the disassembled nanostructures to suppress the background for improving detection sensitivity. Results: As a proof-of-concept study, our findings showed that the application of displacers was more effective in decreasing the SERS intensity of the system and attained a better limit of detection (LOD, 10 aM) than that by directly using activated CRISPR-Cas12a, with high selectivity and stability for nucleic acid detection. Introducing magnetic-responsive functionality to our system further improves the LOD to 1 aM. Conclusion: Our work not only offers a platform to sensitively and selectively probe nucleic acids without pre-amplification but also provides new insights into the design of the CRISPR-Cas12a/SERS integrated system to resolve the steric hindrance of nanomaterials for constructing biosensors.
研究领域:
生物技术
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分析化学
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分子生物学
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DOI
PubMed
高影响因子
Sensing and Bio-Sensing Research
·
2025
·
IF 4.9
Baolin Li,Yaxin Huang,Zixin Zhu,Min Zhong,Guangrong Li,Jinbo Liu
Adenosine triphosphate (ATP) plays a critical role in cellular energy metabolism, acting as the energy source for many biological reactions. Abnormal fluctuations in ATP levels are directly linked to various diseases such as malignancies, bacterial infections, and cardiovascular disorders, making early-stage disease detection through rapid, highly sensitive ATP measurement essential. The current methods for ATP detection suffer from drawbacks such as complex procedures, reliance on expensive instruments, and prolonged reaction times. To this end, we have developed a novel biosensor named ATP Output Sensor Activated by CRISPR (AOSAC).This biosensor employs aptamer-based specific recognition combined with a dual-enzyme, amplification-free system involving Exo III and CRISPR-Cas12a for ATP detection. This sensor operates through an aptamer in a closed duplex structure, upon binding ATP, releases a trigger strand. This strand displaces an activation chain, which in turn interacts with Cas12a's crRNA, initiating trans-cleavage activity that cleaves a single-stranded DNA probe to produce a fluorescent signal. Concurrently, the activation chain's release sites are targeted by Exo III, enhancing signal amplification without the need for complex nucleic acid amplification, thus providing a highly selective, rapid detection method. This technique offers a linear detection range from 0 nM to 20 μM with a detection limit of 44.2 nM, presenting a novel strategy for signal amplification and high specificity in distinguishing ATP from its analogues.
研究领域:
分子生物学
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生物化学
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医学检测
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产品关联
DOI
高影响因子
iScience
·
2024
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IF 4.6
Xuan Wu,Rong Xiang,Danting Yang,Xiaoyao He,Lu Zhu,Fangfang Sun,Hanqing Li,Ningning Pi,Yi Li
Global pandemic has emphasized the needs for advanced pathogen diagnosis in dealing with newly emerged infectious threats, including the Langya henipavirus (LayV). LayV, as an emerging zoonotic pathogen, has potential to cause pandemic, but lacks of rapid diagnostic tools, particularly at point-of-care level. Here, we leveraged the merits of CRISPR-Cas12a biosensing and established a highly sensitive LayV detection method. By integrating CRISPR-Cas12a with RPA, 10 copies/μL ultra-sensitive LayV RNA detection has been achieved at room temperature within 30 min. More importantly, this study developed a special horseradish peroxidase (HRP)-single-stranded DNA (ssDNA) reporter enabling CRISPR-Cas12a to detect LayV RNA without pre-amplification, achieving a sensitivity of 1,200 copies/μL detectable by the naked eye. These explorations can serve as accelerator for CRISPR-Cas biosensing toward rapid response for newly emerged biological threats, and also provide a method to realize simple, precise, and amplicon-free point-of-care pathogen screening for resource limited or underdevelopment regions.
研究领域:
分子生物学
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传染病学
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诊断技术
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产品关联
DOI
PubMed
高影响因子
ANALYTICAL CHEMISTRY
·
2023
·
IF 7.4
Tian Jin Xie,Jia Li Xie,Yu Jie Luo,Kai Mao,Cheng Zhi Huang,Yuan Fang Li,Shu Jun Zhen
DNA nanosheets (DNSs) have been utilized effectively as a fluorescence anisotropy (FA) amplifier for biosensing. But, their sensitivity needs to be further improved. Herein, CRISPR-Cas12a with strong trans-cleavage activity was utilized to enhance the FA amplification ability of DNSs for the sensitive detection of miRNA-155 (miR-155) as a proof-of-principle target. In this method, the hybrid of the recognition probe of miR-155 (T1) and a blocker sequence (T2) was immobilized on the surface of magnetic beads (MBs). In the presence of miR-155, T2 was released by a strand displacement reaction, which activated the trans-cleavage activity of CRISPR-Cas12a. The single-stranded DNA (ssDNA) probe modified with a carboxytetramethylrhodamine (TAMRA) fluorophore was cleaved in large quantities and could not bind to the handle chain on DNSs, inducing a low FA value. In contrast, in the absence of miR-155, T2 could not be released and the trans-cleavage activity of CRISPR-Cas12a could not be activated. The TAMRA-modified ssDNA probe remained intact and was complementary to the handle chain on the DNSs, and a high FA value was obtained. Thus, miR-155 was detected through the obviously decreased FA value with a low limit of detection (LOD) of 40 pM. Impressively, the sensitivity of this method was greatly improved about 322 times by CRISPR-Cas12a, confirming the amazing signal amplification ability of CRISPR-Cas12a. At the same time, the SARS-CoV-2 nucleocapsid protein was detected by the strategy successfully, indicating that this method was general. Moreover, this method has been applied in the analysis of miR-155 in human serum and the lysates of cells, which provides a new avenue for the sensitive determination of biomarkers in biochemical research and disease diagnosis.
研究领域:
生物化学
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分子诊断
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医学检测
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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）