JOURNAL OF PHOTOCHEMISTRY AND PHOTOBIOLOGY A-CHEMISTRY
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2026
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IF 4.7
Li Zhou,Xianwei Jiao,Yuting Zhang,Changling Yan
L-selectin, a leukocyte adhesion receptor, is essential in the early step of the adhesion cascade, facilitating the migration of leukocytes into surrounding tissues during inflammation and immune surveillance. Clinical studies have identified L-selectin as a prospective treatment option for many both acute and ongoing inflammatory conditions. Clarifying the cellular signaling pathways that function in the immediate vicinity of L-selectin may yield unique treatments tailored to certain cell types or illnesses. The study describes the development of a precise and sensitive fluorescence biosensor designed for the detection of L-selectin using a split aptamer specific to L-selectin. This biosensor utilizes nicking enzyme-assisted DNA walking together CRISPR-Cas12a for dual-signal multiplication. The calibration curve for L-selectin demonstrates a detection limit of 0.45 ng/mL, with an extensive linear range spanning from 10 to 1280 ng/mL. The fluorometric sensors effectively identified L-selectin in human blood samples, producing excellent outcomes. The fluorometric sensors effectively identified L-selectin in human blood samples, producing excellent outcomes. The original appearance not only validates the concept but also possesses substantial possibilities for identifying diverse targets.
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TALANTA
·
2026
·
IF 6.7
Mingqi Yang,Jiali Hu,Zhenlin Zhang,Liting Huang,Yahan Yu,Dexing Qiu,Yexin Zu,Yanyan Liu,Zhenyu Lin
Infection with hepatitis B virus (HBV) represents a major challenge to global health, especially in areas of inadequate healthcare infrastructure, rapid and on-site detection plays a crucial role in effective disease management and control. Consequently, it is necessary to develop some simple but sensitive HBV screening techniques. In this work, a homogeneous electrochemical biosensor was constructed for sensitive and specific HBV DNA detection through the integration of CRISPR/Cas12a target recognition and magnetic bead-based separation. The sensor exploits the collateral cleavage activity of CRISPR/Cas12a protein upon recognition of target double-stranded DNA, enabling the degradation of methylene blue-labeled single-stranded DNA captured on magnetic beads (MBs). Then the MBs are separated and redissolved in the solution and the electrochemical response of the system can be tested in homogeneous solution. The electrochemical signal has relationship with the concentration of HBV DNA within a range of 10 fM - 10 nM, and the LOD is 3.74 fM. The sensor demonstrates excellent selectivity and biological stability, which has potential application in clinical diagnosis, especially in resource-limited environments.
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Frontiers in Bioengineering and Biotechnology
·
2025
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IF 4.8
Yunpeng Wu,Jianhua Zheng,Jingqing Chen,Tianqi Sun,Tianqi Luan,Yan Li,Yunzhi Fa,Yefeng Qiu,Rui Zhang
Introduction Human butyrylcholinesterase (hBChE) is a promising bioscavenger against organophosphorus (OP) nerve agents and pesticides. However, low homology-directed repair (HDR) efficiency in CRISPR/Cas9-mediated genome editing limits precise transgene integration in large animals. Methods To improve HDR, we optimized donor structure for targeted integration of hBChE into the goat FGF5 locus using CRISPR/Cas9. Correctly edited goat fibroblast clones were identified by PCR and sequencing. A homozygous clone with reverse-oriented integration was used as a donor for somatic cell nuclear transfer (SCNT), followed by embryo transfer. Offspring were analyzed for genomic integration and transgene expression. Results Reverse-oriented donors significantly enhanced HDR efficiency compared to forward designs, with validation at the pig RAG1 locus. Edited cells stably expressed recombinant hBChE (rhBChE) and showed increased resistance to OP pesticides. SCNT produced a cloned goat expressing high rhBChE levels in the skin. Discussion Optimizing donor structure improves precise genome editing efficiency and enables robust generation of transgenic goats. This strategy advances CRISPR/Cas9-based bioreactor development for scalable production of therapeutic proteins.
研究领域:
遗传学
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分子生物学
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转基因生物技术
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生殖生物学
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ADVANCED FUNCTIONAL MATERIALS
·
2023
·Vol. 33
·期 7
·2210860
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IF 19
Chenya Zhuo,Huiming Kong,Ke Yi,Chun-Wei Chi,Jiabing Zhang,Ran Chen,Haixia Wang,Caixia Wu,Yeh-Hsing Lao,Yu Tao,Mingqiang Li
Chronic hepatitis B infection remains incurable due to the stable presence of various forms of hepatitis B virus (HBV) genome, especially the HBV covalently closed circular DNA (cccDNA). The emergence of clustered regularly interspaced short palindromic repeat (CRISPR) technology provides a new opportunity to potentially cure the HBV infection. However, the efficiency and specificity remain unsatisfactory, especially for nonviral CRISPR/Cas9 delivery. To tackle these, a liver-specific CRISPR/Cas9 magnetic nanosystem FMNP pAG333/sgXPP is constructed based on fluorinated polyethylenimine-coated magnetic nanoparticles and liver-specific ApoE.HCR.hAAT promoter-driven Cas9-T2A-EGFP plasmid with dual sgRNAs. The elaborate system enables magnetic field-induced spatially specific distribution and hepatocyte-specific promoter-driven liver-specific gene editing. Moreover, this CRISPR/Cas9 magnetic nanosystem is designed to disrupt the two conserved sites in X opening reading frame and Pol opening reading frame of the HBV genome, thereby significantly inactivating the HBV genome without showing off-target effects. Treatment with FMNP pAG333/sgXPP for 7 days reduces serum HBsAg levels by 76% with a total editing efficiency of ≈20% in the two conserved sites. Collectively, this study demonstrates spatiotemporal liver genome editing as well as the feasibility of applying a nonviral CRISPR/Cas9 vector for HBV treatment, which may open up a new application for CRISPR therapeutics.
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ANALYTICAL CHEMISTRY
·
2023
·
IF 7.4
Zhirun Ji,Bin Zhou,Zhaoyang Shang,Sirui Liu,Xue Li,Xing Zhang,Bingzhi Li
CRISPR-Cas12a is an accurate and responsive biosensing technique, but its limited stability has restricted its widespread applications. To address this, we propose a strategy using metal–organic frameworks (MOFs) to protect Cas12a from harsh environments. After screening multiple candidate MOFs, it was found that hydrophilic MAF-7 is highly compatible with Cas12a, and the as-formed Cas12a-on-MAF-7 (COM) not only retains high enzymatic activity but also possesses excellent tolerance to heat, salt, and organic solvents. Further investigation showed that COM can serve as an analytical component for nucleic acid detection, resulting in an ultrasensitive assay for SARS-CoV-2 RNA detection with a detection limit of 1 copy. This is the first successful attempt to create an active Cas12a nanobiocomposite that functions as a biosensor without the need for shell deconstruction or enzyme release.
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ANALYST
·
2023
·
IF 4.2
Xinxin Ke,Yi Hu,Chuanxia Chen,Tao Hu
Clustered regularly interspaced short palindromic repeats (CRISPR)-based diagnostics have been considered as the next generation molecular diagnosis tool. Single-readout mode has been extensively employed in massive CRISPR/Cas12a-based biosensors. In this work, we propose a one-tube dual-readout biosensor for ultrasensitive nucleic acids and non-nucleic acids detection by harnessing CRISPR-ALP tandem assay for the first time (termed CRISAT). In the presence of target, the Cas12a is activated to randomly cut the single-stranded hyDNA sequence of MB@hyDNA-cALP, thus releasing abundant alkaline phosphatase (ALP) in the supernatant solution. By using 4-aminophenol phosphate as the substrate of ALP, p-aminophenol is produced and then reacts with N-[3-(trimethoxysilyl)propyl]ethylenediamine or diethylenetriamine to generate silicon-containing polymer carbon dots (Si PCDs) or polymer carbon dots (PCDs) in situ, which can be observed by naked eye or detected by a fluorescent device in the same solution. Using this strategy, a fluoroscence and colorimetry dual-readout nanoplatform for CRISPR-based biosensor can be rationally developed. We ascertain the applicability of CRISAT through detecting SARS-CoV-2 pseudovirus, achieving superior sensitivity and specificity. With simple modification of crRNAs, the CRISAT platform can be also employed to detect monkeypox virus (MPXV) and non-nucleic acids of adenosine triphosphate (ATP). This work provides a great potential for nucleic acids and non-nucleic acids detection.
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ANALYST
·
2022
·Vol. 147
·期 5
·905-914
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IF 5.227
Shumin Zhang,Minyi Liu,Haofa Cui,Muhammad Asad Ziaee,Rongwei Sun,Liting Chen,Daqi Chen,Denis Garoli,Jiahai Wang
The fabrication of nanopores with a matched pore size, and the existence of multiple interferents make the reproducible detection of small-sized molecules by means of solid-state nanopores still challenging. A useful method to solve these problems is based on the detection of large DNA nanostructures related to the existence of small-sized targets. In particular, a DNA tetrahedron with a well-defined 3D nanostructure is the ideal candidate for use as a signal transducer. Here, we demonstrate the detection of an L1-encoding gene of HPV18 as a test DNA target sequence in a reaction buffer solution, where long single-stranded DNA linking DNA tetrahedra onto the surface of the magnetic beads is cleaved by a target DNA-activated CRISPR-cas12 system. The DNA tetrahedra are subsequently released and can be detected by the current pulse in a glassy nanopore. This approach has several advantages: (1) one signal transducer can be used to detect different targets; (2) a glassy nanopore with a pore size much larger than the target DNA fragment can boost the tolerance of the contaminants and interferents which often degrade the performance of a nanopore sensor.
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SENSORS AND ACTUATORS B-CHEMICAL
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2023
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IF 8.4
Jia Wei,Wanjun Wang,Qiong Yu,Mingrui Zhang,Fengyu Xue,Beibei Fan,Tong Zhang,Yanpeng Gao,Juan Li,Xianying Meng,Bo Pang
Mpox (formerly referred as Monkeypox) outbreak has been declared a Public Health Emergency of International Concern. However, traditional polymerase chain reaction (PCR) diagnostic technology is not ideal for on-site applications. To conduct the sample-to-result Mpox viral particles detection outside the laboratories, we developed an easy-to-operate palm-size pouch, termed Mpox At-home Self-Test and point-of-caRe Pouch (MASTR Pouch). In this MASTR Pouch, the fast and accurate visualization was achieved by incorporating recombinase polymerase amplification (RPA) with clustered regularly interspaced short palindromic repeat (CRISPR)/Cas12a system. From viral particle lysis to naked eye readout, MASTR Pouch required only four simple steps to accomplish the analysis process within 35 min. Fifty-three Mpox pseudo-viral particles in exudate (10.6 particles/μL) were able to be detected. To verify the practicability, 104 mock Mpox clinical exudate specimens were tested. The clinical sensitivities were determined to be 91.7%− 95.8%. There was no false-positive result, validating the 100% clinical specificity. MASTR Pouch approaches the WHO's ASSURD criteria for point-of-care diagnostic, which will be beneficial for mitigating Mpox's global spread. The versatility potential of MASTR Pouch could further revolutionize infection diagnosis.
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ANALYTICAL CHEMISTRY
·
2023
·
IF 7.4
Zhenzhen Jia,Yujie Zhang,Chuanyu Zhang,Xueyong Wei,Mingzhen Zhang
With an increasing incidence worldwide, inflammatory bowel disease (IBD) is a chronic inflammatory disease affecting the gastrointestinal tract, which impairs the life quality of patients. Therefore, it is of great significance to construct a sensitive, simple, and convenient biosensor to analyze IBD-associated biomarkers for an auxiliary diagnosis of IBD. Intestinal alkaline phosphatase (IAP), expressed by the intestinal epithelium, is an endogenous protein that is thought to play a vital role in maintaining intestinal homeostasis and is considered a potential biomarker for IBD. Here, an IAP detection method was developed using pregnancy test strips by dephosphorylation. Initially, a double-stranded DNA (dsDNA) was designed to respond to IAP and acted as an activator of Cas12a. In the presence of IAP, the designed dsDNA was not digested by lambda exonuclease (λ exo), which hybridized to the Cas12a-crRNA duplex and resulted in the activation of the trans-cleavage of Cas12a. Further, the activated Cas12a cleaved the single-strand DNA (ssDNA) linker in the MBs-ssDNA-hCG probe, triggering the release of hCG. With magnetic separation, the released hCG could be quantitatively detected by pregnancy test strips. IAP levels were analyzed in feces from colitis and healthy mice by pregnancy test strips. The results showed that the IAP level of colitis mice (3.89 ± 1.92 U/L) was much lower than that of healthy mice (39.64 ± 24.93 U/L), indicating the correlation between IAP and intestinal inflammation. Taken together, a sensitive, user-friendly detection assay based on pregnancy test strips was constructed to monitor IAP and used as an auxiliary diagnostic approach for IBD in a clinical scene.
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TALANTA
·
2021
·
IF 6.057
Xianfeng Wang,Xiaolong Chen,Chengxiang Chu,Yuanyi Deng,Mei Yang,Danqun Huo,Faliang Xu,Changjun Hou,Jiayi Lv
Accurate and effective detection of single-stranded nucleic acids is vital in both disease diagnosis and pathological studies. Hence, we develop a PAMmer-assisted CRISPR/Cas9 system mediated G4-EXPAR (Cas-G4EX) strategy for site-specific detection of ssRNA and ssDNA . PAMmer-assisted CRISPR/Cas9 executes the site-specific cleavage of target ssRNA or ssDNA and released product fragment with the desired sequence at the 3′-terminal. This fragment serves as a primer to activate subsequent sequence-dependent exponential amplification reaction (EXPAR). The G-rich EXPAR products assembles with hemin to form a G-Quadruplex (G4/hemin). G4/hemin catalyzes ABTS-H 2 O 2 system with the appearance of vivid green color, realizing naked-eye analysis. Cas-G4EX integrates the superiority of CRISPR/Cas9 and EXPAR, presenting outstanding site-specific recognition and high-performance amplification efficiency. Meanwhile, the programmability of CRISPR/Cas9 system makes the proposed method become a universal detection paradigm for any ssRNA or ssDNA. Cas-G4EX assay shows the linear relationship from 250 aM to 2.5 nM for ssRNA detection with the actual LOD of 250 aM, and that ranges from 100 aM to 1 nM for ssDNA detection with the actual LOD of 100 aM. Additionally, the acceptable recoveries of 101.48%–109.61% for ssRNA and 93.25%–111.98% for ssDNA in real detection of human serum are obtained for detection of single-strand nucleic acid in real samples. Cas-G4EX also exhibits the excellent discrimination for single-base mutation of single-stranded nucleic acids. Therefore, Cas-G4EX assay provides a promising platform in the applications of molecular diagnosis and pathological analysis.
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BIOSENSORS & BIOELECTRONICS
·
2021
·
IF 10.618
Hui Wu,Yanju Chen,Qunqing Yang,Cheng Peng,Xiaofu Wang,Mengyao Zhang,Siwenjie Qian,Junfeng Xu,Jian Wu
Vibrio parahaemolyticus ( V. parahaemolyticus ) is regarded as a major cause of seafood-associated illnesses, which has aroused widespread public concern. Here, a rapid and convenient detection method for V. parahaemolyticus detection was established by a reversible valve-assisted chip coupling with CRISPR/Cas12a. With optimized lysis buffer, loop mediated isothermal amplification (LAMP) reagents and CRISPR reagents, the whole detection process from sampling to results could be finished within 50 min. The structure of chip was simple and the cost was low. By relying on three reversible rotary valves and the rotation direction-dependent Coriolis pseudo force, the flow order of liquid and the direction of liquid flow could be precisely controlled. The LAMP amplicons were specifically and sensitively identified by CRISPR/Cas12a. Positive amplification would produce green fluorescent signal while negative amplification generated no fluorescent signal, which could be clearly distinguished by the naked eye. With 600 μL of samples processed, the limit of detection (LOD) for both pure cultured V. parahaemolyticus or spiked shrimp samples could achieve 30 copies/reaction. These illustrated the established method displayed great feasibility for real samples detection. In the future, the chip could also combine with other amplification reactions, like PCR or recombinase polymerase amplification reaction (RPA), to conduct detection by changing the corresponding lyophilized amplification reagents. Overall, the proposed detection platform displays great potential for food safety analysis and clinical diagnostics, especially in resource-limited areas.
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Science Advances
·
2019
·Vol. 5
·期 6
·
IF 12.804
Chongyi Liu,Tao Wan,Hui Wang,Song Zhang,Yuan Ping,Yiyun Cheng
Cytosolic protein delivery is of central importance for the development of protein-based biotechnologies and therapeutics; however, efficient intracellular delivery of native proteins remains a challenge. Here, we reported a boronic acid–rich dendrimer with unprecedented efficiency for cytosolic delivery of native proteins. The dendrimer could bind with both negatively and positively charged proteins and efficiently delivered 13 cargo proteins into the cytosol of living cells. All the delivered proteins kept their bioactivities after cytosolic delivery. The dendrimer ensures efficient intracellular delivery of Cas9 protein into various cell lines and showed high efficiency in CRISPR-Cas9 genome editing. The rationally designed boronic acid–rich dendrimer permits the development of an efficient platform with high generality for the delivery of native proteins.
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