One-step Operation Research Articles

In-situ nucleic acid amplification induced by DNA self-assembly for rapid and ultrasensitive detection of miRNA

BackgroundTo improve the sensitivity and specificity of nucleic acid detection, coupling two or more signal amplification systems is a feasible pattern, such as nucleic acid isothermal amplification coupling genome-editing technology, and cascaded DNA self-assembly circuits. And representative signal amplification strategies include loop-mediated isothermal amplification (LAMP), clustered regularly interspaced short palindromic repeats/associated proteins (CRISPR/Cas) systems, and catalyzed hairpin assembly (CHA). However, these detection strategies often require the enrichment of intermediate products, the replacement of reaction conditions and the design of multiple probes, which may seriously affect the reliability of detection results. ResultsHerein, we propose a novel nucleic acid detection system which is named as catalyzed hairpin assembly (CHA) coupled with embedded primer triggered isothermal amplification (CEA for short). DNA self-assembly probes in CEA contain a specially designed primer. When target nucleic acid (e.g., miRNA) initiates CHA reaction (the first signal amplification), the self-assembly product of CHA will expose a primer (named as embedded primer). The embedded primer will trigger a special nucleic acid isothermal amplification in situ, then generate plenty of double-stranded DNA products in 30 min with varying lengths (the second signal amplification). Compared to that of a typical CHA reaction, the sensitivity of CEA has increased by three orders of magnitude and the detection limit is as low as 0.228 fM. Besides, it has excellent detection performance in cancer and stem cell samples. SignificanceBy coupling embedded primer with DNA self-assembly system, a new nucleic acid detection system (CEA) with one-step operation and dual signal amplification has been successfully established. Compared with traditional dual signal amplification systems, CEA can not only significantly improve the reaction efficiency, but also greatly reduce the difficulty of detection system design and experimental operation.

Mix-and-Read Digital MicroRNA Analysis Based on Flow Cytometric Counting of Target-Clicked Nanobead Dimer.

A one-step, enzyme-free, and highly sensitive digital microRNA (miRNA) assay is rationally devised based on flow cytometric counting of target miRNA-clicked nanobead dimers via a facile mix-and-read manner. In this strategy, highly efficient miRNA-sandwiched click chemical ligation of two DNA probes may remarkably stabilize and boost the dimer formation between two kinds of fluorescence-coded nanobeads, and the number of as-produced bead dimers will be target dose-responsive, particularly when the trace number of miRNA is much less than that of employed nanobeads. Finally, each fluorescence-coded bead dimer can be easily identified and digitally counted by a powerful flow cytometer (FCM) and accordingly, the amount of target miRNA can be accurately quantified in a digital way. This new digital miRNA assay can be accomplished with a facile mix-and-read operation just by simply mixing the target miRNA with two kinds of preprepared DNA probe-functionalized nanobeads, which do not require any nucleic acid amplification, purification, and complex operation procedures. In spite of the extremely simple one-step operation, benefiting from the low-background but high target-mediated click ligation efficiency, and the powerfully digital statistical capability of FCM, this strategy achieves high sensitivity with a quite low detection limit of 5.2 fM target miRNA as well as high specificity and good generality for miRNA analysis, pioneering a new direction for fabricating digital bioassays.

The Outcome of a Single Stage Repair for Proximal Hypospadias in Otherwise Healthy Child at Mymensingh Medical College Hospital

Hypospadias is a birth defect that involves the development of a penile urethra that opens on the ventral side of the penis. Proximal hypospadias is one of the most severe forms of hypospadias that is generally associated with chordee. Hypospadias surgery, especially in the proximal location, is challenging, and many operations have been proposed, including one-stage and two-stage operations. Objectives: The aim of the study was to evaluate the success rate and safety of a single-stage surgery in children with proximal hypospadias and to determine the complications. Materials and Methods: This interventional study was conducted on forty patients, aged between one and twelve years, who had proximal hypospadias and were admitted to Mymensingh Medical College Hospital between January 2014 and April 2015. A one-step operation with tabularized incised plate urethroplasty (TIPU) was done. Patients’ demographic data, intraoperative details, and postoperative outcomes were recorded within 3 months after the operation. Results: The common age at surgery was 2-4 years, and they were 40% of the total patients. There was Haematoma in 5% of cases, a devitalized skin flap in 7.5% of cases, and stent blockage in 10% of cases during the immediate postoperative period. In our 3-month follow-up, all the patients had conical glans and cylindrical penises. Meatal stenosis and narrow stream were noted in 5 percent of patients but were relieved with meatal dilatation. Urethrocutaneous fistulas remained patients in 7.5% of cases. Conclusion: TIPU repair of proximal hypospadias is safe, effective, yields good results, and has low risks of complications in otherwise healthy children. It is a less risky and more viable method that helps to exclude extra steps in the process.

One-step time-resolved cascade logic gate microfluidic chip for home testing of SARS-CoV-2 and flu B

Home testing technology strategy is critical for early screening of disease. However, current home testing technologies often require complex processes, which limits their application. In this study, a time-resolved cascade logic gate microfluidic chip (TCLMC) was revealed to enable capillary force-based one-step operation without manual intervention or professional equipment. By analogy with logic gates in the circuit, TCLMC could automatically control the fluid flow and regulate the incubation time to optimize the immunoassay. The limit of detection of TCLMC for SARS-CoV-2 and influenza B virus (Flu B) was 134.94 and 79.17 pg mL−1 within 10 min. Additionally, this study tested saliva samples from 12 Flu B patients and 24 healthy controls to verify its clinical application. The results showed that TCLMC had high sensitivity (100%), specificity (100%), and accuracy (100%). This study provides a new one-step strategy for home testing and demonstrates its great potential in the diagnosis field.

Scalable one-step synthesis of reduced graphene oxide: Towards flexible transparent conductive films and active supercapacitor electrodes

The reduced graphene oxide (rGO) thin films have attracted much attention in flexible electronic devices and supercapacitors (SCs). However, the chemical reduction mechanism of GO needs to be clarified, and the one-step process at low temperatures for preparing high-quality rGO thin films from GO dispersions is still blank in the research field. Here, we propose a one-step operation to spray GO dispersions onto substrate or current collector under a hydroiodic acid (HI) atmosphere while simultaneously achieving the preparation of rGO thin films. This pioneering metal-free, transfer-free, fast, low-temperature, environmentally friendly, and scalable spraying technology enables large-scale production of high-quality rGO. This method realizes the preparation of highly conductive and high transmittance rGO thin films on flexible polyimide and polyethylene terephthalate substrates, as well as the preparation of rGO thin films on current collectors as SCs electrodes. Density functional theory calculations show that oxygen-containing functional groups on the surface of GO exhibit a stronger adsorption energy for HI, and have a low energy barrier during reduction by HI. Ab initio molecular dynamics simulated the possible reduction reaction path between GO and HI at 90 °C, demonstrating that HI molecules can effectively reduce GO. Furthermore, the sheet resistance of the transparent conductive film created using the method developed in this study reaches approximately 1000 Ω/sq (80 % transparency), and the specific capacitance of the SCs electrode film created in this method is 433.2 F/g (5 mV/s, 1 M H2SO4). Our study not only validates the efficacy of the one-step synthesis method for rGO thin films but also provides theoretical insights into the underlying reaction mechanism. This work holds significant promise for advancing applications in various electronics by facilitating large-scale production of high-quality rGO thin films.

An ultrasensitive and multiplexed miRNA one-step real time RT-qPCR detection system and its application in esophageal cancer serum

MicroRNAs (miRNAs) are increasingly recognized as promising biomarkers for early disease diagnosis and prognosis. Therefore, the need for rapid, robust methods for multiplex miRNA detection in biological research and clinical diagnosis is crucial. This study introduces a novel multiplex miRNA detection method, SMOS-qPCR (Sensitive and Multiplexed One-Step RT-qPCR). The method integrates multiplexed reverse transcription and TaqMan-based qPCR into a single tube, employing a one-step operation on a real-time PCR system. We investigated the effect of 3′ end phosphorylation of the Linker, Linker concentration and probe concentration on the SMOS-qPCR, resulted in a wide linear range from 1 fM to 0.1 zM (R2 ≥ 0.99 for each miRNA), surpassing the capabilities of stem-loop RT-qPCR and SYBR Green One-step RT-qPCR. The method showed excellent performance in distinguishing mature miRNA from miRNA precursor, and successfully detected four miRNAs in a single tube without cross-interference. Its high specificity enables precise differentiation of less than 1% nonspecific signal. Finally, we demonstrated the effectiveness of the SMOS-qPCR system in detecting circulating miRNAs in serum samples, distinguishing between esophageal cancers and health individuals with high AUC values (>0.940). In conclusion, the proposed SMOS-qPCR system offers a straightforward and promising approach for miRNA profiling in future clinical applications.

Double-Cross-Linked Hydrogel with Long-Lasting Underwater Adhesion: Enhancement of Maxillofacial In Situ and Onlay Bone Retention.

Bone retention is a usual clinical problem existing in a lot of maxillofacial surgeries involving bone reconstruction and bone transplantation, which puts forward the requirements for bone adhesives that are stable, durable, biosafe, and biodegradable in wet environment. To relieve the suffering of patients during maxillofacial surgery with one-step operation and satisfying repair, herein, we developed a double-cross-linked A-O hydrogel named by its two components: [(3-Aminopropyl) methacrylamide]-co-{[Tris(hydroxymethyl) methyl] acrylamide} and oxidated methylcellulose. With excellent bone adhesion ability, it can maintain long-lasting stable underwater bone adhesion for over 14 days, holding a maximum adhesion strength of 2.32 MPa. Schiff-base reaction and high-density hydrogen bonds endow the hydrogel with strong cohesion and adhesion performance as well as maneuverable properties such as easy formation and injectability. A-O hydrogel not only presents rarely reported long-lasting underwater adhesion of hard tissue but also owns inherent biocompatibility and biodegradation properties with a porous structure that facilitates the survival of bone graft. Compared to the commercial cyanoacrylate adhesive (3 M Vetbond Tissue Adhesive), the A-O hydrogel is confirmed to be safer, more stable, and more effective in calvarial in situ bone retention model and onlay bone retention model of rat, providing a practical solution for the everyday scenario of clinical bone retention.

UV-induced graft polymerization of polyamide-6 for electroless copper deposition

A simple and chromium-free method for the metallization on engineering plastic polyamide-6 (PA6) by UV irradiation is proposed in this article. The surface graft polymers with specific adsorption activity are developed through one-step UV exposure operation, which captures Pd2+ in the activation process. The chemisorption of Pd2+ is further reduced into catalytic Pd0 to enable the substrate with catalytic nucleus for conventional copper electroless deposition (ELD). The grafting layer and the deposited copper coating are characterized by Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, contact angle measurements, scanning electron microscopy, and X-ray diffraction. The results indicate that successful grafting occurs on PA6 after UV excitation. In addition, the as-deposited copper layer shows excellent conductive performance and high adhesion with the PA6 substrate. It’s expected that this emerging and feasible approach would have broad prospects for the metallization of plastic and offer significant value in the plating industry.

One ethane-selective metal–organic framework with customized pore size and specific binding sites for efficient purification of ethylene

Removing small amount of ethane (C2H6) from ethylene (C2H4) to obtain high-purity C2H4 is of great significance in the industries. However, the construction of C2H6-selective metal–organic frameworks (MOFs) remains challenging, mainly because that C2H4 has smaller kinetic diameter and larger quadrupole moment. Herein, an ultra-microporous MOF (PCP-bdc) with customized pore size and specific binding sites is reported to efficiently purify C2H4 from C2H6/C2H4 mixture. The channel size of PCP-bdc is comparable to the size of C2H6 molecule, which can provide more accessible channel surfaces for C2H6. The channel surface of PCP-bdc is rich of low-polar aromatic groups and the uncoordinated O atoms can interact with C2H6 molecules strongly. Thus, PCP-bdc exhibits a remarkable C2H6/C2H4 (50/50) selectivity (2.82) and C2H6 uptake (53.8 cm3 g−1) at 298 K and 1 bar, which outperform those of many C2H6-selective adsorbents. Density functional theory calculations further reveal the formation of multiple synergies between this MOF and C2H6 enables the efficient separation for C2H6/C2H4 mixture. Breakthrough experiments demonstrate that PCP-bdc can purify C2H4 from C2H6/C2H4 mixture through one-step operation. These results indicate PCP-bdc is a promising adsorbent for C2H6/C2H4 separation.

Continuous enantiomeric separation using water-oil-water segmented flow system

The multiphase segmented flow, which consisted of the material, extraction, and recovery phases, was applied to the chiral separation. The use of simultaneous extraction and back-extraction with multiple phases aims to prevent the accumulation of target material in the extraction phase by efficiently removing it in the recovery phase, thereby enabling extraction performance beyond the equilibrium. This study demonstrated the chiral separation of an aqueous racemic amino acid derivative (3,5-dinitrobenzoyl-(R,S)-leucine) with a cinchona alkaloid chiral host. A droplet manipulation system comprising PFA manifolds and valves was utilized to alternately form an acidic material phase slug and a basic recovery phase slug in a 1-octanol oil phase containing the chiral host. Two aqueous phases were collected separately using valves controlled by an optical sensor signal. The manipulation system achieved stable operation across various conditions, including flow rate, residence time, and slug lengths. The enantiomeric excess, which represents the purity of the chirality, could be increased to 50 % through 200 s extraction in a one-step operation.

Cascade capture, oxidization and inactivation for removing multi-species pollutants, antimicrobial resistance and pathogenicity from hospital wastewater

As reservoirs of pathogens, antimicrobial resistant microorganisms and a wide variety of pollutants, hospital wastewaters (HWWs) need to be effectively treated before discharge. This study employed the functionalized colloidal microbubble technology as one-step fast HWW treatment. Inorganic coagulant (monomeric Fe(III)-coagulant or polymeric Al(III)-coagulant) and ozone were used as surface-decorator and gaseous core modifier, respectively. The Fe(III)- or Al(III)-modified colloidal gas (or, ozone) microbubbles (Fe(III)-CCGMBs, Fe(III)-CCOMBs, Al(III)-CCGMBs and Al(III)-CCOMBs) were constructed. Within 3 min, CCOMBs decreased CODCr and fecal coliform concentration to the levels meeting the national discharge standard for medical organization. Regrowth of bacteria was inhibited and biodegradability of organics was increased after the simultaneous oxidation and cell-inactivation process. The metagenomics analysis further reveals that Al(III)-CCOMBs performed best in capturing the virulence genes, antibiotic resistance genes and their potential hosts. The horizontal transfer of those harmful genes could be effectively hampered thanks to the removal of mobile genetic elements. Interestingly, the virulence factors of adherence, micronutrient uptake/acquisition and phase invasion could facilitate the interface-dominated capture. Featured as cascade processes of capture, oxidation and inactivation in the one-step operation, the robust Al(III)-CCOMB treatment is recommended for the HWW treatment and the protection of downstream aquatic environment.

A ratiometric fluorescence platform for lead ion detection via RNA cleavage-inhibited self-assembly of three-arm branched junction

Heavy metal pollution can pose a threat to food safety and human health, and accurate quantification of heavy metal ions is a vital requirement. Emerging DNA nanostructures-based biosensors offer attractive tools toward ultra-sensitive or rapid analysis of heavy metal ions. However, the problems including complex design, severe reaction conditions and undesirable reliability are inevitable obstacle in advancing their extension and application. Herein, a ratiometric fluorescent platform was established for monitoring lead ion (Pb2+) in food based on dual Förster resonance energy transfer (FRET) and RNA cleavage-inhibited self-assembly of three-arm branched junction (TBJ). GR-5 DNAzyme was employed for Pb2+ recognition, and enzyme-free amplification technique catalytic hairpin assembly (CHA) served to form FRET probes-carried TBJ. The substrate strand (S) of DNAzyme triggered the generation of CHA-TBJ, and Pb2+-responsive cleavage of S hindered the assembly of CHA-TBJ, causing opposite changes in the FRET states of FAM/BHQ1 and ROX/BHQ2 pairs. The fluorescence responses were recorded through synchronous fluorescence spectrometry to indicate Pb2+ concentration, allowing sensitive and reliable identification of Pb2+ in the linear range of 0.05–5 ng mL−1 with the detection limit of 0.03 ng mL−1. The Pb2+ detection can be achieved under conventional reaction conditions, simple mixing procedures and one-step measurement operation. The approach can afford excellent specificity for Pb2+ against competing metal ions, and can be applied to analyze Pb2+ in tea samples with satisfactory results. This facile fluorescence platform shows a capable method for Pb2+ detection, and provides new avenue in the development of ratiometric approaches and DNAzyme strategies for monitoring heavy metal pollution, facilitating the transformation of DNAzyme-based biosensors for food safety control.

One-step generation of Bell state on nonlocal acoustics wave resonators assisted by nitrogen-vacancy center ensembles

Recently, quantum information processing (QIP) on acoustics wave resonators (AWRs) has attracted much attention as the quality factor of AWR has been increased to 1011, which means the time of phonons stored in the AWR can reach the order of seconds. To achieve the large-scale QIP on AWRs, one should complete quantum entangled operations on nonlocal AWRs. Different from previous work, we propose a one-step all-resonance scheme to generate Bell states on two nonlocal AWRs coupled to two nitrogen-vacancy center ensembles (linked by an AWR quantum bus) respectively. One-step all-resonance operation makes the scheme easier to be experimentally implemented.

One-Step Fabrication of Droplet Arrays Using a Biomimetic Structural Chip.

In the field of one-step efficient preparation of dewetting droplet arrays, the process is hampered by the requirement for low chemical wettability of solid surfaces, which restricts the complete transition of wetting state and its broad prospects in biological applications. Inspired by the physical structure of the lotus leaf, enabling it to promote the change of the infiltration state of an aqueous solution on the surface, we developed a method of one-step fabrication of droplet arrays on the biomimetic structural chip designed in the present work. This greatly reduces the need for chemical modification techniques to achieve low wettability and reduces the reliance on complex and sophisticated surface preparation techniques, thus improving the fabrication efficiency of droplet arrays fully generated on a chip by one-step operation without the need for extra liquid phase or the control of harsh barometric pressure. We also studied the influence of dimensions of the biomimetic structure and the preparation process parameters such as number of smears and speed of smearing on the preparation rate and uniformity of the droplet arrays. The amplification of templating DNA molecules in the droplet arrays prepared in a one-step fabrication way is also performed to verify its application potential for DNA molecular diagnosis.

Selective extraction of thorium to directly form self-assembly solid from HNO3 solution

Based on ions exchange between [DMDSA]+[Cl]- (Dimethyl distearyl ammonium chloride) and N,N-dialkyl-succinamide acid (SCA), three novel bifunctional [DMDSA]+[SCA]- ionic liquids (ILs) were firstly synthesized for extraction of thorium (IV) by self-assembly strategy. The simultaneous extraction and solidification of Th(IV) were unexpectedly realized in one-step operation using the present ILs in HNO3 solution, and more than 99% thorium (IV) was enriched and immediately aggregated into self-assembly solid at the biphasic interface. The self-assembly solid was further identified by FT-IR, SEM with element mapping EDS and XPS analysis, and revealing that the self-assembly extraction (SAE) was triggered by the amphiphilic [DMDSA]+ cations. A three-step extraction mechanism dominated by [SCA·Th(NO3)4]- was proposed based on the slope analysis method and HRMS analysis. The self-assembly extraction of Th(IV) exhibited the extremely excellent selectivity in the presence of U(VI) and typical lanthanide elements including La(III), Eu(III) and Lu(III), and the separation factors reached 2516 for Th/U, 1885 for Th/La, 1512 for Th/Eu and 558 for Th/Lu, respectively. The proposed SAE strategy was proved to be an efficient method for one-step separation and solidification of thorium ions from U(VI) and/or lanthanides.

Wheel drive-based DNA sensing system for highly specific and rapid one-step detection of MiRNAs at the attomolar level

With the use of DNA as building blocks, a variety of microRNA amplification-based sensing systems have been developed. Nevertheless, ultrasensitive, selective and rapid detection of microRNAs with a high signal-to-background ratio and point mutation discrimination ability remains a challenge. Herein, we propose a novel wheel drive-based DNA sensing system (NWDS) based on a self-assembled, self-quenched nanoprobe (SQP) to conduct highly specific and ultrasensitive one-step measurement of microRNAs. In this work, a signalling recognition DNA hairpin (DH) sequence with a self-complementary stem domain of 14 base pairs was used, which contained three functional regions, namely a recognition region for the target miRNA-21, a sticky region with 9 complementary nucleotides to the 3′terminus of a DNA wheel (DW) and a region for the hybridization with a quenching DNA primer (DP). The SQP was ingeniously self-assembled at room temperature by the DH and DP, which was capable of eliminating unwanted background signals. MiRNA-21 was employed as a target model to specifically activate the SQP, leading to specific hybridization between the HP and DW. With the assistance of a polymerase, an SQP-based wheel driving took place to induce hybridization/polymerization displacement cycles, initiating target recycling and DP displacement. As a result, a large amount of the newly formed hybrid SQP/DW accumulated to generate a substantially enhanced fluorescence signal. In this way, the newly proposed NWDS exhibits ultrasensitivity with a detection limit of 5.62 aM across a wide linear dynamic response range up to 200 nM, excellent selectivity with the capability to discriminate homologous miRNAs and one-base, two-base and three-base mismatched sequences, and an outstanding analytical performance in complex systems. In addition, the significant simultaneous advantages of one-step operation, rapid detection within 15 min and a high signal-to-background ratio of 26 offer a unique opportunity to promote the early diagnosis of cancer-related diseases and molecular biological analysis.

Simultaneous detection of ultraviolet irradiation and vitamin C using an all-carbon-based integrated wearable system powering by a micro-supercapacitor

Ultraviolet (UV) radiation is a harmful exogenous factor for human skin. Wearable UV photodetectors can monitor UV exposure in the surroundings, and wearable vitamin C (VC) sensors tracking the levels in the human body present the potential ability to defend the UV radiation. Herein, we reported on the fabrication of an all-in-one wearable system with a UV photodetector and VC sensor powered by a micro-supercapacitor. Based on direct laser writing carbonization of polyimide sheets, the patterned electrodes and interconnects of the circuit were fabricated by a facile one-step operation, obtaining an all-carbon-based integrated system. Such a system exhibited outstanding energy storage ability (56.2 μWh cm−2 at 4.17 mW cm−2), high areal capacitance (1.06 mF cm−2 at 5 mV s−1), satisfying capacitive stability, and good mechanical flexibility. The UV photodetector and the VC sensor were powered to obtain a linear range of UV intensity from 11 to 44 μW cm−2 (equivalent to Ultraviolet Index 4.4 to 17.6), and VC levels of 1.0–200 μM with a low limit of detection of 0.83 μM. Furthermore, the integrated system was successfully applied to the determination of VC in commercial beverage and human sweat samples. This work provided a simple and promising method to fabricate integrated wearable systems for on-site providing information on the UV intensity of the external environment and the VC level of the human body simultaneously.

Two colors, one-step, self-drive fluorescent strategy for chloramphenicol detection base on DNAzyme cleavage triggered hybridization chain reaction

A two colors, one-step, self-drive fluorescent strategy was developed for chloramphenicol (CAP) detection based on cyclic cleavage of molecular beacon (MB) by pincer DNA sequences. CAP can bind with its aptamer and active the enzyme-strand (E-DNA). Then the E-DNA can circularly cleave the MB on the both side of pincer DNA sequences. The cleaved fragments can self-assembly to form a long duplex and cause the great recovery of the two colors fluorescent signal. The limit of detection was as low as 0.7 pM. Importantly, the whole detection process is very simple with only one-step operation. Moreover, the two colors fluorescent signals can greatly enhance the accuracy of the result. It was also successfully used to detect CAP in actual samples.

Polyhalogenation-Facilitated Spirolactonization at the meta-Position of Phenols

A novel dearomative spirolactonization/polyhalogenation of phenols that employs hypervalent iodine PhICl2 (iodobenzene dichloride) as both an oxidant and chlorine source with an indispensable base, or only using NBS (N-bromosuccinimide) without any additives, is presented. Halide participations are a vital factor in the cascade reaction of 3'-hydroxy-[1,1'-biphenyl]-2-carboxylic acids with good selectivities and reactivities and induced the rapid constructions of multiple C-halogen bonds and directional C═O bonds in a one-step operation under mild conditions. In gaining a good understanding of the mechanism, the increase in number of bromine atoms was inferred rationally from the spirolactonization process, assisted by DFT calculations and high-resolution mass spectrometry. Mechanistic experiments suggest that the formation of a stable carbocation intermediate plays a great role in the migration of oxygen to spirolactonization.

Continuous-flow macromolecular sieving in slanted nanofilter array: stochastic model and coupling effect of electrostatic and steric hindrance.

Microfabricated slanted nanofilter arrays are a promising technology for integrated biomolecule analysis systems such as online monitoring and point-of-care quality validation, due to their continuous-flow and one-step operation capability. However, an incomplete understanding of the system limits the performance and wider applications of slanted nanofilter arrays. In this paper, we present rigorous theoretical and experimental studies on macromolecule sieving in a slanted nanofilter array. From both stochastic and kinetic models, an explicit theoretical solution describing size-dependent molecule sieving was derived, which was validated using experimental sieving results obtained for various sieving conditions. Our results not only detail the relationship between sieving conditions and sieving efficiency but also demonstrate that sieving is affected by multiple hindrance effects (electrostatic hindrance), not steric hindrance alone. There is an optimal sieving condition for achieving the greatest separation efficiency for DNAs of a certain size range. Small DNA has great size selectivity in small nanofilters and in weak electric fields, whereas large DNA is present in large nanofilters and in strong electric fields. This study provides insights into designing a slanted nanofilter array for particular target applications and understanding the sieving principles in the nanofilter array.