Enhanced CRISPR/Cas12a Fluorimetry via a DNAzyme-Embedded Framework Nucleic Acid Substrate.

CRISPR/Cas12a fluorimetry has been extensively developed in the biosensing arena, on account of its high selectivity, simplicity, and rapidness. However, typical CRISPR/Cas12a fluorimetry suffers from low sensitivity due to the limited trans-cleavage efficiency of Cas12a, necessitating the integration of other preamplification techniques. Herein, we develop an enhanced CRISPR/Cas12a fluorimetry via a DNAzyme-embedded framework nucleic acid (FNAzyme) substrate, which was designed by embedding four CLICK-17 DNAzymes into a rigid tetrahedral scaffold. FNAzyme can not only enhance the trans-cleavage efficiency of CRISPR/Cas12a by facilitating the exposure of trans-substrate to Cas12a but also result in an exceptionally high signal-to-noise ratio by mediating enzymatic click reaction. Combined with a functional nucleic acid recognition module, this method can profile methicillin-resistant Staphylococcus aureus as low as 18 CFU/mL, whose sensitivity is approximately 54-fold higher than that of TaqMan probe-mediated CRISPR/Cas12a fluorimetry. Meanwhile, the method exhibited satisfactory recoveries in food matrices ranging from 80% to 101%. The DNA extraction- and preamplification-free detection format as well as the potent detection performance highlight its tremendous potential as a next-generation analysis tool.

Artificial Intelligence-Assisted Digital Immunoassay Based on a Programmable-Particle-Decoding Technique for Multitarget Ultrasensitive Detection.

The development of a multitarget ultrasensitive immunoassay is significant to fields such as medical research, clinical diagnosis, and food safety inspection. In this study, an artificial intelligence (AI)-assisted programmable-particle-decoding technique (APT)-based digital immunoassay system was developed to perform multitarget ultrasensitive detection. Multitarget was encoded by programmable polystyrene (PS) microspheres with different characteristics (particle size and number), and subsequent visible signals were recorded under an optical microscope after the immune reaction. The resultant images were further analyzed using a customized, AI-based computer vision technique to decode the intrinsic properties of polystyrene microspheres and to reveal the types and concentrations of targets. Our strategy has successfully detected multiple inflammatory markers in clinical serum and antibiotics with a broad detection range from pg/mL to µg/mL without extra signal amplification and conversion. An AI-based digital immunoassay system exhibits great potential to be used for the next generation of multitarget detection in disease screening for candidate patients.

Magnetic Relaxation Switching Immunoassay Based on Hydrogen Peroxide-Mediated Assembly of Ag@Au-Fe3 O4 Nanoprobe for Detection of Aflatoxin B1.

Magnetic relaxation switching (MRS) sensors have shown great potential in food safety monitoring due to their high signal-to-noise ratio and simplicity, but they often suffer from insufficient sensitivity and stability due to the lack of excellent magnetic nanoprobes. Herein, dumbbell-like Au-Fe3 O4 nanoparticles are designed as magnetic nanoprobes for developing an aflatoxin B1-MRS immunosensor. The Fe3 O4 portion in the Au-Fe3 O4 nanoparticles functions as the magnetic probe to provide transverse relaxation signals, while the Au segments serve as a bridge to grow Ag shell and assemble the Au-Fe3 O4 nanoparticles, thus modulating transverse relaxation time of surrounding water molecular. The formation of Ag@Au-Fe3 O4 is triggered by hydrogen peroxide. After degraded by horseradish peroxidase, hydrogen peroxide reduces Ag+ to Ag nanoparticles which assemble dispersed Au-Fe3 O4 to aggregated Ag@Au-Fe3 O4 , thus dramatically improving the sensitivity of traditional MRS sensor. Combined with competitive immunoreaction, this Ag@Au-Fe3 O4 -MRS immunosensor can detect aflatoxin B1 with a high sensitivity (3.81 pg mL-1 ), which improved about 21 folds and 9 folds than those of enzyme-linked immunosorbent assay and high-performance liquid chromatography (HPLC), respectively. The good consistency with HPLC in real samples detection indicates the good accuracy of this immunosensor. This Ag@Au-Fe3 O4 -MRS immunosensor offers an attractive tool for detection of harmful substances.

Development of Cu(II)/Cu(I)-induced quantum dot-mediated fluorescence immunoassay for the sensitive determination of ethyl carbamate.

A series of haptens were rationally designed for producing monoclonal antibodies specific for EC and a simple fluorescence immunoassay platform was developed for the sensitive determination of EC based on alkaline phosphatase (ALP)-triggered Cu+ quenching of CdSe quantum dots (QDs). It was noted that Cd as a fluorescence substrate in CdSe QDs can be selectively substituted by Cu+ that resulted in a more significant fluorescence quenching in comparison with Cu2+. Meanwhile, because ALP catalyzed ascorbic acid phosphate and then assisted the transformation of Cu2+ to Cu+, the change in fluorescence intensity was found to be proportional to ALP concentration. After simple magnetic separation, the sensitivity and linear range of the established assay were improved approximately 53-fold and an order of magnitude, respectively, when compared with the conventional ELISA. The proposed platform was able to both amplify the signal and eliminate matrix interferences, making it a promising to determine EC as well as other contaminants in complex food matrix in a highly sensitive and simple manner. Graphical abstract.

Amplified Magnetic Resonance Sensing via Enzyme-Mediated Click Chemistry and Magnetic Separation.

Magnetic resonance sensing (MRS) assays with simple pretreatment have drawn increasing attention for the development of biosensors, whereas conventional MRS is not competent for analyzing trace targets due to its relative low sensitivity. In this work, we employ alkaline phosphatase-mediated click chemistry as a signal transformation and amplification technique that effectively enhances the magnetic signal for amplified MRS. We outline two strategies by modulating either the aggregation state or the number of the magnetic probes based on click chemistry-triggered aggregation of nanoparticles and their ability for magnetic separation. The experimental results show that the combination of magnetic nanoprobe number-based modulation and magnetic separation outcompetes nanoprobe aggregation-based strategy for an enhanced sensitivity, providing an attractive platform for MRS-based analysis of small molecules. For proof of application, we apply this click chemistry-based approach for the detection of clenbuterol, an illegally used drug to promote growth in livestock. It shows a linear detection range from 0.1 ng mL-1 to 500 ng mL-1 with a limit of detection of 0.02 ng mL-1, revealing that the click chemistry-mediated amplified MRS can act as a promising magnetic platform to detect trace levels of small molecules in authentic samples.

Cu(II)-induced magnetic resonance tuning and enhanced magnetic relaxation switching immunosensor for sensitive detection of chlorpyrifos and Salmonella.

Magnetic relaxation switching (MRS) biosensors have been recognized as useful analytical tools for a range of targets; however, traditional MRS biosensors are limited by the "prozone effect", resulting in a narrow linear range and low sensitivity. Herein, we proposed a paramagnetic Cu2+-induced magnetic resonance tuning (MRET) strategy, based on which Cu2+ ions and magnetic nanoparticles (MNPs) were adopted to construct a Cu-MNP-mediated MRS (Cu-M-MRS) immunosensor with Cu2+ ions acting as a quencher and MNPs as an enhancer. An Fe3O4@polydopamine-secondary antibody conjugate was prepared and used to correlate the amount of Cu2+ ions to the target concentration through an immunoassay. Based on the immunoreaction, the Cu-M-MRS immunosensor enabled the sensitive detection of chlorpyrifos (0.05 ng/mL, a 77-fold enhancement in sensitivity compared with the traditional MRS immunosensor) and Salmonella (50 CFU/mL). The proposed MRET strategy effectively improved the sensitivity and accuracy of the MRS immunosensor, offering a promising and versatile platform for food safety detection.

CRISPR/Cas12a and Hybridization Chain Reaction-Coregulated Magnetic Relaxation Switching Biosensor for Sensitive Detection of Viable Salmonella in Animal-Derived Foods.

We combined a CRISPR/Cas12a system with a hybridization chain reaction (HCR) to develop an ultrasensitive magnetic relaxation switching (MRS) biosensor for detecting viable Salmonella typhimurium (S. typhimurium). Magnetic nanoparticles of two sizes (30 and 1000 nm: MNP30 and MNP1000, respectively) were coupled through HCR. The S. typhimurium gene-activated CRISPR/Cas12a system released MNP30 from the MNP1000-HCR-MNP30 complex through a trans-cleavage reaction. After magnetic separation, released MNP30 was collected from the supernatant and served as a transverse relaxation time (T2) signal probe. Quantitative detection of S. typhimurium is achieved by establishing a linear relationship between the change in T2 and the target gene. The biosensor's limit of detection was 77 CFU/mL (LOD = 3S/M, S = 22.30, M = 0.87), and the linear range was 102-108 CFU/mL. The accuracy for detecting S. typhimurium in real samples is comparable to that of qPCR. Thus, this is a promising method for the rapid and effective detection of foodborne pathogens.

Magnetic relaxation switching immunosensor based on polystyrene microcolumn and tyramine signal amplification for ultrasensitive and user-friendly detection of aflatoxin B1 in corn.

Aflatoxin B1 is highly mutagenic in humans, and long-term exposure can impair immunity and increase the risk of cancer. It is imperative to develop immunoassays with convenient operation and high sensitivity to detect aflatoxin B1. This study presents a polystyrene microcolumn-mediated magnetic relaxation switching immunosensor based on a tyramine signal amplification strategy for detecting aflatoxin B1. An environmentally friendly hand-held polystyrene microcolumn was designed as an effective immunoreaction carrier, remaining 91% efficiency after 12 repeated uses. And the microcolumn provides a user-friendly procedure for rapid separation and reagent switching within 3 s by simple stirring in solution. The combination of a strong anti-interference magnetic relaxation switching biosensing and an efficient tyramine signal amplification enables the quantitative detection of aflatoxin B1 in the range of 0.01-10 ng/mL, with a limit of detection of 0.006 ng/mL. This method has potential application in the rapid detection of trace food contaminants.

Multienzyme-Mediated Dual-Channel Magnetic Relaxation Switching Taste Biosensor (D-MRSTB) for Simultaneous Detection of Umami Compounds and Synergistic Enhancement in Food.

Umami substances play a significant role in the evaluation of food quality, and their synergistic enhancement is of great importance in improving and intensifying food flavors and tastes. Current biosensors available for umami detection still confront challenges in simultaneous quantification of multiple umami substances and umami intensities. In this study, an innovative dual-channel magnetic relaxation switching taste biosensor (D-MRSTB) was developed for the quantitative detection of representative umami substances. The multienzyme signal of D-MRSTB specifically catalyzes the umami substances of interest to generate hydrogen peroxide (H2O2), which is then used to oxidate Fe2+ to Fe3+. Such a valence-state transition of paramagnetic ions was utilized as a magnetic relaxation signaling switch to influence the transverse magnetic relaxation time (T2) within the reaction milieu, thus achieving simultaneous detection of monosodium glutamate (MSG) and inosine 5'-monophosphate (IMP). The biosensor showed good linearity (R2 > 0.99) in the concentration range of 50-1000 and 10-1000 µmol/L, with limits of detection (LOD) of 0.61 and 0.09 µmol/L for MSG and IMP, respectively. Furthermore, the biosensor accurately characterized the synergistic effect of the mixed solution of IMP and MSG, where ΔT2 showed a good linear relationship with the equivalent umami concentration (EUC) of the mixed solution (R2 = 0.998). Moreover, the D-MRSTB successfully achieved the quantitative detection of umami compounds in real samples. This sensing technology provides a powerful tool for achieving the detection of synergistic enhancement among umami compounds and demonstrates its potential for application in the food industry.

Experimental Investigation of Reflectarray Antennas for High-Power Microwave Applications.

The power capacity of reflectarray antennas (RAs) is investigated through full-wave simulations and high-power microwave (HPM) experiments in this paper. In order to illustrate the results in detail, two RA elements are designed. The simulated power handling capacity of two RA elements are 7.17 MW/m2 and 2.3 GW/m2, respectively. To further study the HPM RA, two RA prototypes operating at 2.8 GHz are constructed with the aperture size of 1 m × 1 m. Simulations and experimental measurements are conducted for the two prototypes. The experimental results demonstrate that, even when subjected to 1 GW of power, the radiation beam of the RA with the second elements can still propagate in the intended direction. This research will establish a basis for advancing the practicality of RAs in HPM applications.

A machine vision-assisted Argonaute-mediated fluorescence biosensor for the detection of viable Salmonella in food without convoluted DNA extraction and amplification procedures.

The precise identification viable pathogens hold paramount significance in the prevention of foodborne diseases outbreaks. In this study, we integrated machine vision and learning with single microsphere to develop a phage and Clostridium butyricum Argonaute (CbAgo)-mediated fluorescence biosensor for detecting viable Salmonella typhimurium (S. typhimurium) without convoluted DNA extraction and amplification procedures. Phage and lysis buffer was utilized to capture and lyse viable S. typhimurium, respectively. Subsequently, CbAgo can cleave the bacterial DNA to obtain target DNA that guides a newly targeted cleavage of fluorescent probes. After that, the resulting fluorescent signal accumulates on the streptavidin-modified single microsphere. The overall detection process is then analyzed and interpreted by machine vision and learning algorithms, achieving highly sensitive detection of S. typhimurium with a limit of detection at 40.5 CFU/mL and a linear range of 50-107 CFU/mL. Furthermore, the proposed biosensor demonstrates standard recovery rates and coefficients of variation at 93.22% - 106.02% and 1.47% - 12.75%, respectively. This biosensor exhibits exceptional sensitivity and selectivity, presenting a promising method for the rapid and effective detection of foodborne pathogens. ENVIRONMENTAL IMPLICATION: Bacterial pathogens exist widely in the environment and seriously threaten the safety of human life. In this study, we developed a phage and Clostridium butyricum Argonaute-mediated fluorescence biosensor for the detection of viable Salmonella typhimurium in environmental water and food samples. Compared with other Salmonella detection methods, this method does not need complex DNA extraction and amplification steps, which reduces the use of chemical reagents and experimental consumables in classic DNA extraction kit methods.

DNA Extraction- and Amplification-Free Nucleic Acid Biosensor for the Detection of Foodborne Pathogens Based on CRISPR/Cas12a and Argonaute Protein-Mediated Cascade Signal Amplification.

A novel method for detecting low levels of viable foodborne pathogens, specifically Salmonella typhimurium (S. typhimurium), has been developed. Traditional nucleic acid assay, such as polymerase chain reaction (PCR), often requires complex DNA extraction and amplification, making it challenging to differentiate between viable and nonviable pathogens. This assay employed a phage as the recognition element to precisely identify and lyse viable S. typhimurium that can undergo DNA extraction. It combined the efficient trans-cleavage activities of CRISPR/Cas12a with the specific cleavage advantages of Argonaute proteins, enabling ultrasensitive detection. This double-enzyme-mediated nucleic acid test can accurately distinguish viable and nonviable S. typhimurium with a detection limit of 23 CFU/mL without DNA amplification. The method was successfully applied to common food samples, producing results consistent with quantitative PCR tests. This work provides a promising platform for easily detecting viable foodborne pathogens with high sensitivity without the need for DNA extraction and amplification.

Computer vision-assisted smartphone microscope imaging digital immunosensor based on click chemistry-mediated microsphere counting technology for the detection of aflatoxin B1 in peanuts.

Aflatoxin B1 is a carcinogenic contaminant in food or feed, and it poses a serious health risk to humans. Herein, a computer vision-assisted smartphone microscope imaging digital (SMID) immunosensor based on the click chemistry-mediated microsphere counting technology was designed for the detection of aflatoxin B1 in peanuts. In this SMID immunosensor, the modified polystyrene (PS) microspheres were used as the signal probes and were recorded by a smartphone microscopic imaging system after immunoreaction and click chemistry reaction. The number of PS probes is adjusted by aflatoxin B1. The customized computer vision procedure was used to efficiently identify and count the obtained PS probes. This SMID immunosensor enables sensitive detection of aflatoxin B1 with a linear range from 0.001 ng/mL to 500 ng/mL, providing a simple, sensitive, and portable tool for food safety supervision.

Magnetic relaxation switching immunoassay based on "limited-magnitude" particles for sensitive quantification of aflatoxin B1.

Aflatoxin B1 (AFB1) is a highly toxic and carcinogenic chemical substance that endangers food safety and human health. Magnetic relaxation switching (MRS) immunosensors are utilized in a variety of applications in food analysis due to its resistance to matrix interferences, but they often suffer from magnetic separation-based multi-washing steps and low sensitivity. Herein, we propose novel MRS strategy for the sensitive detection of AFB1 using "Limited-Magnitude" size particles: a single millimeter sized polystyrene spheres (PSmm) and 150 nm superparamagnetic nanoparticles (MNP150). Only a single PSmm is used as the microreactor to enhance all of the magnetic signal on its surface in high concentration by an immune competitive response, successfully preventing signal dilution, which can be transferred by pipette, simplifying the process of separation and washing. The established single polystyrene sphere magnetic relaxation switch biosensor (SMRS) was able to quantify AFB1 from 0.02 to 200 ng/mL with a detection limit of 14.3 pg/mL. SMRS biosensor has been successfully used for the detection of AFB1 in wheat and maize samples, and the results in agreement with high performance liquid chromatography-tandem mass spectrometry (HPLC-MS). Benefiting from high sensitivity and convenient operation, the simple and enzyme-free method is promising in trace small molecules applications.

Computer Vision-Based Artificial Intelligence-Mediated Encoding-Decoding for Multiplexed Microfluidic Digital Immunoassay.

Digital immunoassays with multiplexed capacity, ultrahigh sensitivity, and broad affordability are urgently required in clinical diagnosis, food safety, and environmental monitoring. In this work, a multidimensional digital immunoassay has been developed through microparticle-based encoding and artificial intelligence-based decoding, enabling multiplexed detection with high sensitivity and convenient operation. The information encoded in the features of microspheres, including their size, number, and color, allows for the simultaneous identification and accurate quantification of multiple targets. Computer vision-based artificial intelligence can analyze the microscopy images for information decoding and output identification results visually. Moreover, the optical microscopy imaging can be well integrated with the microfluidic platform, allowing for encoding-decoding through the computer vision-based artificial intelligence. This microfluidic digital immunoassay can simultaneously analyze multiple inflammatory markers and antibiotics within 30 min with high sensitivity and a broad detection range from pg/mL to µg/mL, which holds great promise as an intelligent bioassay for next-generation multiplexed biosensing.

Magnetic Relaxation Switching Immunosensors via a Click Chemistry-Mediated Controllable Aggregation Strategy for Direct Detection of Chlorpyrifos.

Chlorpyrifos (CPF) is the most frequently found organophosphate pesticide residue in solid food samples and can cause increasing public concerns about potential risks to human health. Traditional detection signals of such small molecules are mostly generated by target-mediated indirect conversion, which tends to be detrimental to sensitivity and accuracy. Herein, a novel magnetic relaxation switching detection platform was developed for target-mediated direct and sensitive detection of CPF with a controllable aggregation strategy based on a bioorthogonal ligation reaction between tetrazine (Tz) and trans-cyclooctene (TCO) ligands. Under optimal conditions, this sensor can achieve a detection limit of 37 pg/mL with a broad linear range of 0.1-500 ng/mL in 45 min, which is approximately 51-fold lower than that of the gas chromatography analysis and 13-fold lower than that of the enzyme-linked immunosorbent assay. The proposed click chemistry-mediated controllable aggregation strategy is direct, rapid, and sensitive, indicating great potential for residue screening in food matrices.

Cartridge voltage-sensitive micropump immunosensor based on a self-assembled polydopamine coating mediated signal amplification strategy.

Current biosensing detection assays were developed to focus on rapid, low-cost, stable detection for clinical diagnosis and food safety monitoring. In this work, a novel portable cartridge voltage-sensitive micropump immunosensor (CVMS) biosensing device based on the integration of the microchannel circuit biosensing principle and polydopamine (PDA) was presented for rapid and sensitive detection of pathogenic factors in real samples at trace levels. The CVMS can sensitively evaluate voltage signal changes caused by clogging effects in the closed-loop circuit when the insulated microspheres pass through the microchannel. The targets could trigger the immune reaction between antibody-antigens that leads to the change in the concentration of horseradish peroxidase (HRP). And the HRP was further employed to catalyze the polymerization of dopamine into PDA, resulting in the rapid formation of the magnetic @PDA nanoparticles (MNP@PDA) with core-shell structures. The abundant functional groups on the MNP@PDA surface can efficiently adsorb polystyrene microspheres, thus causing changes in the number of polystyrene microspheres (PS). The CVMS can accurately monitor the change of PS with a portable device, which weighs less than 0.8 kg and costs only $50. The completion of CVMS takes 90 min to complete. The limit of detection of this immunosensor for procalcitonin and ochratoxin A were 42 pg/mL and 77 pg/mL, respectively, which improved about 15 folds and 38 folds, respectively, than those of enzyme-linked immunosorbent assay.

Single-Layer Wide-Angle Scanning Linear Phased Arrays Based on Multimode Microstrip Patch Elements.

This paper introduces a novel single-layer microstrip patch element designed to achieve a wide beamwidth, in order to address the growing demand for wide-angle scanning capabilities in modern phased array systems. The proposed element, comprising a slot-etched circular patch and an array of metallized holes arranged in square rings, offers a unique approach to beam shaping. By carefully adjusting parameters such as the slot structure and feeding position, our element is engineered to simultaneously excite both the TM01 and TM21 modes, a key feature that contributes to its wide beamwidth characteristics. Through the constructive interference of these modes, our element demonstrates a remarkable 3 dB beamwidth of approximately 150° in both principal planes, showcasing its potential for wide-angle scanning applications. To validate the practical performance of this proposed element, two linear phased arrays are manufactured and experimentally evaluated. The simulation results confirm the wide-angle scanning capability of the antennas in both the E-plane and H-plane. Furthermore, the experimental assessment demonstrates that these linear phased arrays can effectively generate scanning beams within a frequency range of 25 GHz to 28 GHz, covering a wide angular range from -60° to 60°, while maintaining a gain loss within 3 dB. This innovative design approach not only offers a promising solution for achieving a wide beamwidth in microstrip patch elements, but also holds significant potential for the development of cost-effective phased arrays with wide-angle scanning capabilities, making it a valuable contribution to the advancement of phased array technology.

Magnetic relaxation switching biosensor via polydopamine nanoparticle mediated click chemistry for detection of chlorpyrifos.

Traditional magnetic relaxation switching (MRS) biosensors suffer from poor sensitivity and unsatisfactory stability. In this study, a polydopamine (PDA) nanoparticles (NPs)-Cu2+ chelate complex mediated signal conversion system and a Cu+-catalyzed click chemistry triggered magnetic signal amplification system were evaluated and dynamically integrated into an MRS biosensor. Owing to abundant functional groups and a large surface area, PDA NPs enabled the absorption of a large amount of Cu2+ ions by chelation. The residual Cu2+ ions can be reduced with sodium ascorbate to Cu+, which could initiate the click reaction between azide-functionalized magnetic NPs (MNPs) and alkyne-functionalized MNPs that resulted in the production of aggregated nanoclusters. The transverse relaxation time (T2) depends on the degree of aggregation of MNPs; T2 is expressed as the magnetic signal readout. In addition, PDA NPs can be easily conjugated with antibodies by mixing, thus providing a straightforward bridge that integrates the immunoassay and magnetic signal readout. Combined with the high capacity of PDA NPs for chelating Cu2+ and high efficiency of click reaction for changing the T2 signals, the PDA-MRS biosensor enables the detection of chlorpyrifos with a limit of detection of 0.084 ng/mL, providing 22-fold enhancement than traditional enzyme-linked immunosorbent assay (1.86 ng/mL). This demonstrates its great potential for the detection of hazardous chemical molecules in a complex sample matrix.

[Separation and enrichment of trace aflatoxin B1 in grains by magnetic nanomaterials based on SiO2@Fe3O4].

In this study, a magnetic nanomaterial antibody (Ab)-SiO2@Fe3O4 was synthesized, which was employed to absorb aflatoxin B1 (AFB1) in complicated grain matrices. The Ab-SiO2@Fe3O4 material was then paired with high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) for subsequent accurate detection. The Ab-SiO2@Fe3O4 material has a specific adsorption capacity for AFB1 because of the stable and specific biological binding between antigen and antibody. This process can achieve the identification between the material and food matrix quickly, thereby completing the separation and enrichment process. Then, high sensitivity and high accuracy HPLC-MS/MS were employed for signal readout and actual quantification, which can significantly increase the detection efficiency and enable high-throughput detection of numerous samples. In the pretreatment process, Fe3O4 was first synthesized by microwave-assisted hydrothermal synthesis within 1 h, and Ab-SiO2@Fe3O4 was then produced using the enhanced Stober's approach. This material with high adsorption performance was synthesized under relatively mild conditions and short time. To obtain Ab-SiO2@Fe3O4 materials with uniform particle size, magnetic properties, and dispersibility that met the requirements, synthesis conditions of Ab-SiO2@Fe3O4 and conditions for capturing the AFB1 target were analyzed. The findings demonstrated that the best effect was obtained when the dosage of FeCl3·6H2O was 10.0 mmol, the heating time was 40 min, and 100 µL tetraethoxysilane was employed for SiO2 coating. The AFB1 antibody was then combined with the surface of SiO2@Fe3O4 under several conditions. The findings revealed that the best coupling efficiency of Ab could be obtained when the concentration of 2-morpholinoethanesulfonic acid monohydrate (MES) was 10 mmol/L, pH was 6.5, and the molar ratio of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC)∶N-hydroxysuccinimide substances (NHS) was 2∶1. The coupling buffer was then selected as phosphate buffer (PBS) with pH=7.4, and 8 mg Ab-SiO2@Fe3O4 was employed to separate and enrich AFB1 at 37 ℃ for 10 min. In the actual detection, acetonitrile-water-formic acid (85∶10∶5, v/v/v) was employed as the extraction solution. After ultrasonic extraction for 10 min, Ab-SiO2@Fe3O4 was employed to separate and enrich AFB1 in the extract. The supernatant was dried with nitrogen and reconstituted with 1-mL acetonitrile. The solution was then filtered through a 0.22 µm filter and detected using HPLC-MS/MS, thereby realizing the quick and quantitative detection of AFB1. AFB1 had an excellent linear relationship in the range of 2-50 µg/L under the optimal analytical conditions, and the correlation coefficient was less than 0.99. The LOD was 0.04 µg/kg, and the LOQ was 0.13 µg/kg. The spiked recoveries of AFB1 in three grain matrices ranged from 76.21% to 92.85% with RSD≤5.29% at four different spiked levels. The approach was applied to the determination and analysis of AFB1 in 30 real grain samples of rice, corn, and wheat. The findings demonstrated that AFB1 was detected in one wheat sample and two corn samples, and its content was 0.38, 0.13, and 0.47 µg/kg, respectively, and no toxins were found in other samples. The approach combined Ab-SiO2@Fe3O4 magnetic nanomaterials with HPLC-MS/MS, which could obtain high-efficiency separation and enrichment of AFB1. Furthermore, the low-cost Ab-SiO2@Fe3O4 could be stored for more than a week and complete the pretreatment process within 30 min. This effective pretreatment process combined with HPLC-MS/MS could realize the analysis of several samples within a short time, and had a promising application prospect in the detection of AFB1 in grains.