Nonspecific Adsorption Research Articles

Identification of Antibody to Low-Frequency Antigen in Thalassemia: Non-specific Allo Adsorption Method

Identification of Antibody to Low-Frequency Antigen in Thalassemia: Non-specific Allo Adsorption Method

Development of Functional Biointerface Using Mixed Zwitterionic Silatranes.

Strategies to design multifunctional interfaces for biosensors have been extensively investigated to acquire optimal sensitivity, specificity, and accuracy. However, heterogeneous ingredients in clinical samples inevitably generate background signals, exposing challenges in biosensor performance. Polymer coating has been recognized as a crucial method to functionalize biointerfaces by providing tailored properties that are essential for interacting with biological systems. Herein, we introduce for the first time two oligomeric silatranes, MPS-MPCn and MPS-PEGMACOOHm, which were copolymerized from mercaptopropylsilatrane (MPS) with either zwitterionic monomer 2-methacryloyloxyethyl phosphorylcholine (MPC) or carboxylated poly(ethylene glycol) methacrylate (PEGMACOOH) through thiol-ene polymerization. These oligomeric silatranes were prepared individually and in combinations in acidic and nonacid solvents for deposition on silicon wafers. Afterward, coating properties, including wettability, thickness, and elemental composition, were characterized by contact angle meter, ellipsometer, and X-ray photoelectron spectroscopy (XPS), respectively. Importantly, MPS-MPCn polymers were found to form thin films with high hydrophilicity and superior fouling repulsion to bacteria and protein, while mixed coating involving 70% MPS-PEGMACOOH2.5 and 30% MPS-MPC2.5 exhibited thinnest coating with best wettability among COOH-terminated coatings. Furthermore, the functional COOH group in the coated surfaces was exploited for postmodification with biological molecules via intermediated N-hydroxysuccinimide (NHS) ester group by amine coupling chemistry. Once again, the combination of 70% MPS-PEGMACOOH2.5 and 30% MPS-MPC2.5 provided an ultimate reduction in nonspecific adsorption (NSA) and established a finest signal discrimination through enzyme-linked immunosorbent assay. Consequently, these novel mixed oligomeric silatranes offer a promising approach for the construction of biosensor interfaces with dual functions in both nonspecific binding prevention and conjugation of biomolecules.

Highly Sensitive and Biocompatible Microsensor for Selective Dynamic Monitoring of Dopamine in Rat Brain.

Highly selective and sensitive in vivo neurotransmitter dynamic monitoring of the central nervous system has long been a challenging endeavor. Here, an implantable and biocompatible microsensor with excellent performances was reported by electrodepositing poly(3,4-ethylenedioxythiophene)-electrochemically reduced graphene oxide (PEDOT-ERGO) nanocomposites and poly(tannic acid) (pTA) sequentially on the carbon fiber electrode (CFE) surface, and its feasibility in in vivo electrochemical sensing applications were demonstrated. Due to the synergistic electrocatalytic effect of PEDOT-ERGO nanocomposites with the negative-charged pTA on dopamine (DA) redox reaction, the microsensor exhibits high detection sensitivities of 1.1 and 0.37 nA μM-1 in the detection ranges of 0.02-0.5 and 0.5-20 μM with a low limit of detection of 9.2 nM. Also, the microsensor shows excellent selectivity, good sensing stability, repeatability, and reproducibility. In addition, the highly hydrophilic and negative-charged pTA inhibits the nonspecific adsorption of hydrophobic proteins, which endows the microsensor with good antifouling ability. Moreover, DA dynamics in rat brain were successfully monitored in real time, and the selective sensing ability of the microsensor in vivo was also demonstrated. The present study provides a new method for selective dynamics monitoring of DA in the brain, which would help to better understand the pathological and physiological functions of DA.

Bifunctional MoO3-x/CuS Heterojunction Nanozyme-Driven "Turn-On" SERS Signal for the Sensitive Detection of Cerebral Infarction Biomarker S100B.

The use of nanozymes has become a promising auxiliary approach to "turn on" surface-enhanced Raman scattering (SERS) signals for the label-free detection of disease markers. Nevertheless, there are still major challenges to develop bifunctional nanomaterials with both excellent enzyme-like activity and high SERS performance. To this end, a novel Z-scheme MoO3-x/CuS heterojunction was first constructed as a powerful "two-in-one" substrate, which can not only catalyze leucomalachite green (LMG) to SERS-active malachite green (MG) but also serve as an efficient substrate to amplify the SERS signal of catalysate. Due to the strong interfacial coupling effect between the MoO3-x and CuS nanomaterial, which promoted the separation and transport of carriers in the heterojunction, the MoO3-x/CuS heterojunction showed higher peroxidase-like activity compared to individual components and the previously reported heterojunction nanozymes. Inspired by these results, a sandwich-type SERS immunoassay for the detection of the cerebral infarction biomarker S100 calcium-binding protein (S100B) was proposed based on the output signal of MG at 1620 cm-1. Furthermore, introducing the antifouling material chitosan on the surface of the MoO3-x/CuS heterojunction can effectively resist nonspecific protein adsorption and significantly improve the detection accuracy of the immunoassay. Therefore, the SERS immunoassay based on the MoO3-x/CuS heterojunction realized highly sensitive and selective detection of S100B in the concentration range of 0.001 to 100 ng/mL, with a low limit of detection of 0.47 pg/mL. The developed method has been successfully used for the accurate detection of S100B in clinical serum. The results showed that the level of S100B in the serum of cerebral infarction patients can be distinguished from those of healthy individuals and intracranial tumor patient controls. In addition, the acquired values of S100B in the serum of cerebral infarction patients based this strategy were well consistent with the results of electrochemiluminescence (ECL) detection with a relative error of less than ±7.3. It is expected that this work may open up a paradigm for improving detection sensitivity and accuracy for the early diagnosis and treatment monitoring of cerebral infarction in the clinic.

Zwitterionic Polymers for Biomedical Applications: Antimicrobial and Antifouling Strategies toward Implantable Medical Devices and Drug Delivery.

Poly(ethylene glycol) (PEG) is extensively utilized in biomedical applications due to its biocompatibility; however, its thermal instability and susceptibility to oxidative degradation significantly constrain its long-term effectiveness. Zwitterionic polymers, characterized by their distinctive structure, enhanced stability, and superior biocompatibility, offer a more advantageous alternative. These polymers exhibit super hydrophilicity, resist nonspecific protein adsorption, and maintain stability in biological environments due to their charge-neutral ionic nature. Zwitterionic polymers enhance anticancer drug delivery by precisely targeting tumor cells and facilitating an efficient drug release. Their inherent antifouling properties and prolonged circulation within the bloodstream render them highly suitable for redox-sensitive drug carriers, thereby augmenting the antitumor efficacy. Moreover, zwitterionic polymers markedly mitigate biofouling in implants, biosensors, and wound dressings, thereby improving both their functionality and their therapeutic outcomes. These advantages arise from the formation of robust hydration layers, which significantly enhance the hemocompatibility and inhibit the adhesion of proteins, platelets, and bacteria. Zwitterionic polymers, including sulfobetaine (SB), phosphorylcholine (PC), and carboxybetaine (CB), are increasingly employed in blood-contacting devices and as effective coating materials for implantable devices. This mini-review paper aims to explore the recent diverse biomedical applications of zwitterionic polymers and highlight their advantageous properties compared with unmodified polymers. We will cover their use in drug delivery systems, tumor targeting nanocarriers, antibiofouling and antibacterial activities in implantable devices, tissue engineering, and diagnostic devices, demonstrating how their unique properties can translate into different applications. Through this exploration, this Perspective will display the potential of zwitterionic polymers as innovative polymer materials in the field of biomedical engineering and beyond.

Single-Extracellular-Vesicle Detection with a Plasmonic Chip and Enhanced Fluorescence Microscopy.

Endocytosis-derived extracellular vesicles (EVs), which can be as small as 100 nm, are useful for disease prediction. However, very small EVs are below the optical diffraction limit and are difficult to visualize with conventional fluorescence microscopy. In this study, single EVs captured on a plasmonic chip, where fluorescently labeled antibodies were bound over the EV surface, were detected as bright spots using plasmon-field enhanced fluorescence without any pretreatment of isolating labeled EVs, followed by analyzing the full width at half-maximum and the fluorescence peak value for each enhanced fluorescence bright spot. Bright spots smaller than the threshold determined by the observation of the fluorescent nanospheres were attributed to single EVs. The number of single EVs was quantitatively evaluated against the concentration of EV solution injected in the 1.4 pM-95 fM range. Furthermore, single EVs were detected by labeling two different membrane proteins. A molecularly imprinted polymer was applied to a capture interface on a plasmonic chip, and it is found that nonspecific adsorption of aggregates was suppressed. To accurately distinguish single EVs from aggregates of labeled antibodies, the fluorescence microscopy with transmitted light was superior to the epifluorescence method. Finally, single EVs were successfully detected with multiple targets at multiple wavelengths by using different fluorescently labeled antibodies.

Acoustic pipette and biofunctional elastomeric microparticle system for rapid picomolar-level biomolecule detection in whole blood.

Most biosensing techniques require complex processing steps that generate prolonged workflows and introduce potential points of error. Here, we report an acoustic pipette to purify and label biomarkers in 70 minutes. A key aspect of this technology is the use of functional negative acoustic contrast particles (fNACPs), which display biorecognition motifs for the specific capture of biomarkers from whole blood. Because of their large size and compressibility, the fNACPs robustly trap along the pressure antinodes of a standing wave and separate from blood components in under 60 seconds with >99% efficiency. fNACPs are subsequently fluorescently labeled in the pipette and are analyzed by both a custom, portable fluorimeter and flow cytometer. We demonstrate the detection of anti-ovalbumin antibodies from blood at picomolar levels (35 to 60 pM) with integrated controls showing minimal nonspecific adsorption. Overall, this system offers a simple and versatile approach for the rapid, sensitive, and specific capture of biomolecules.

HaloTag-Modified, Ferrocene Labeled Self-Assembled Monolayers for Protein Sensing.

Ferrocene (Fc)-based disulfide molecules of various lengths with amino acid scaffolds and alkane or oligo(phenylene-ethynylene) (OPE) bridges are used in a mixed SAM with a di-(ethylene oxide) terminal mercaptoundecanol diluent (PEG2). The relative height of the Fc redox reporter in the SAM is compared to determine if there are protective effects like antifouling and specific detection. The HaloTag-binding motif is used as a proof-of-concept to investigate the electrochemical response to the HaloTag protein due to its known covalent and fast linkage. When the Fc-SAMs are exposed to the HaloTag protein, there are an antifouling nature and more specific detection for the engulfed Fc-based molecules (C6tBu/Halo). The further out the Fc is from the SAM layer, the more nonspecific adsorption is detected. The double layer capacitance (CDL) has the smallest change for the C6tBu control (ΔCDL = -0.1 μF cm-2) showing antifouling properties and produces a large change (ΔCDL = 0.9 μF cm-2) as well as a shift in oxidation potential when the active C6Halo is exposed to the HaloTag protein (ΔE1/2 = 50 ± 10 mV). The remaining Fc molecules are partially in or outside the PEG2 layer, allowing more ion penetration/mobility even when the HaloTag protein is bound. Generally, a more disordered environment was observed for the Fc-based molecules when adding the HaloTag ligand, which is evident from a larger Efwhm and higher CDL. Desorption of the SAMs with sodium iodide (NaI) showed retention of the HaloTag protein bound with the corresponding ligand, whereas negative controls did not. Self-assembled monolayers for MALDI mass spectrometry (SAMDI-MS) were used as an orthogonal detection technique to show the qualitative binding of the HaloTag protein to the electrode. Together, these results provide insight into the antifouling and detection methods of engulfing the redox molecules in the SAM diluent.

Phosphate-Driven Interfacial Self-Assembly of Silk Fibroin for Continuous Noncovalent Growth of Nanothin Defect-Free Coatings.

Silk fibroin is a fiber-forming protein derived from the thread of Bombyx mori silkworm cocoons. This biocompatible protein, under the kosmotropic influence of potassium phosphate, can undergo supramolecular self-assembly driven by a random coil to β-sheet secondary structure transition. By leveraging concurrent nonspecific adsorption and self-assembly of silk fibroin, we demonstrate an interfacial phenomenon that yields adherent, defect-free nanothin protein coatings that grow continuously in time, without observable saturation in mass deposition. This noncovalent growth of silk fibroin coatings is a departure from traditionally studied protein adsorption phenomena, which generally yield adsorbed layers that saturate in mass with time and often do not completely cover the surface. Here, we explore the fundamental mechanisms of coating growth by examining the effects of coating solution parameters that promote or inhibit silk fibroin self-assembly. Results show a strong dependence of coating kinetics and structure on solution pH, salt species, and salt concentration. Moreover, coating growth was observed to occur in two stages: an early stage driven by protein-surface interactions and a late stage driven by protein-protein interactions. To describe this phenomenon, we developed a kinetic adsorption model with Langmuir-like behavior at early times and a constant steady-state growth rate at later times. Structural analysis by FTIR and photoinduced force microscopy show that small β-sheet-rich structures serve as anchoring sites for absorbing protein nanoaggregates, which is critical for coating formation. Additionally, β-sheets are preferentially located at the interface between protein nanoaggregates in the coating, suggesting their role in forming stable, robust coatings.

Adsorption of Proteins onto Nitrocellulose Membranes from a Flowing Solution – Theory and Experiment

Some analytical laboratory procedures involve passing the sample through a porous polymer membrane. In this process, the analyte binds to the surface of the membrane modified with a specific receptor layer and is then detected using optical or electrochemical signals. This paper presents an experimental and theoretical analysis of the binding patterns of the analyte to nitrocellulose membranes. Two cases are considered: specific binding of the analyte to antibodies immobilized on the membrane and non-specific adsorption of the analyte. It is shown that increasing the volume of the sample passed through the membrane leads to an increase in the amount of adsorbed analyte, and this can generally be used to improve the sensitivity of biosensors.

Manganese nanoparticles catalyzed the formation of mesoporous helical nitrogen-doped carbon nanotubes enabled aldosterone biosensing

Primary aldosteronism (PA) is the foremost form of secondary hypertension that affects 5–20 % of healthy humans via the excessive secretion of aldosterone (ALD), which increases the level of potassium excretion and thus produces various critical combined diseases. Thus, determining trace-level ALD content in human blood or urine samples is a crucial factor in diagnosing a life-threatening disease called PA. Herein, nanosized manganese particles encapsulated within the tunable helical-shaped N-doped mesoporous carbon nanotubes (Mnx@H-NCNTs) are synthesized by a simple and facile ball-milling method at different reaction times (1, 6, 12, and 18 h). The influence of ball-milling process time on the final morphology of the as-synthesized Mnx@H-NCNTs nanostructures is systematically investigated. In order to fabricate an ALD biosensor electrode, the immobilization of Anti-ALD antibodies (Anti-ALD) on the surface of the Mn@H-NCNTs-12H electrode (Anti-ALD/Mn@H-NCNTs-12H) is achieved via the interactions of combined interactions of hydrogen bonds, Van der Waals’ force, negatively charged carboxylic acid functionalities of antibodies and positively charged metal centers, and π-π stacking interaction between nucleosides of Anti-ALD and mesoporous H-NCNTs. In addition, BSA was used as a blocking additive for covering non-specific adsorption sites of Anti-ALD/Mnx@H-NCNTs (BSA/Anti-ALD/Mn@H-NCNTs-12H) nanostructured electrodes. The BSA/Anti-ALD/Mn@H-NCNTs-12H can be used as an effective signal amplifying probe for ALD detection through a noticeable reduction in the amperometric i-t current response over a linear range from 1 to 2500 pM, with a LOD of 0.72 pM (S/N = 3). The practical applicability of BSA/Anti-ALD/Mn@H-NCNTs-12H is further demonstrated by the detection of ALD from 1 to 20 nM in human blood or urine samples with a recovery ranging from 90.5 to 97.0 %, indicating its potential use in clinical analysis.

Hydrogen-Bonded Organic Framework Enhanced Antifouling Property for Efficient In Situ Electrochemical Assay of Cerebral Ascorbic Acid.

Accurate determination of cerebral ascorbic acid (AA) is crucial for understanding ischemic stroke (IS) related pathological events. Carbon fiber microelectrodes (CFEs) have proven to be robust tools with high sensitivity toward AA, however, they face ongoing challenges for in situ measurement due to the non-specific adsorption of proteins in brain tissue. In this study, the hydrogen-bonded organic framework PFC-71 is synthesized and modified on CFEs through π-π stacking interactions with carboxylated carbon nanotubes (CNT-COOH). It is found that the gating effect and hydrophilicity of PFC-71 provided the CFE with excellent antibiofouling properties. As a result, AA exhibited a low oxidation potential of -30mV on the CFE/CNT-COOH/PFC-71, even in the presence of 20mgmL-1 bovine serum albumin. Given the structural advantages of CFE/CNT-COOH/PFC-71, a ratiometric electrochemical strategy for AA is established, enabling the in situ assay of cerebral AA in a middle cerebral artery occlusion (MCAO) model with high accuracy and stability.

Zwitterionic Peptides: From Mechanism, Design Strategies to Applications.

Zwitterionic peptides, as a type of peptide composed of charged residues, are electrically neutral, which combine the advantages of zwitterionic materials and biological peptides, exhibiting hydrophilicity and programmable properties. As attractive candidates for resisting nonspecific adsorption of biomacromolecules and microorganisms, zwitterionic peptides have been applied in materials science, biomedicine, and biochemistry over the past decade. In this review, the development of zwitterionic peptides has been systematically outlined and analyzed, including their mechanisms, structure-function relationships, and design strategies. Furthermore, this review emphasizes and discusses their recent applications for developing functional coatings, biosensors, drug delivery systems, and engineering proteins. Finally, future research perspectives and challenges of zwitterionic peptides are also prospected and discussed. This review is intended to provide clarity and insight into the design and applications of zwitterionic peptides.

Dual responsive drug-loaded nanomotor based on zwitterionic materials for the treatment of peritoneal metastatic cancer

Innovative treatments for peritoneal metastatic cancer have attracted widespread attention from researchers. Here, we propose a drug-loaded nanomotor (PSBMA/l-Arg/DOX, PLD) based on zwitterionic materials for the treatment of peritoneal metastatic cancer through intraperitoneal injection. Zwitterionic polymer nanocarriers (PSBMA NPs) are obtained by radical polymerization with zwitterionic SBMA as the polymerization monomer and N,N’-Bis(acryloyl)cystamine (BAC) as the cross-linking agent. The zwitterionic substrate of this nanomotor has the ability to resist non-specific protein adsorption in ascites. The loaded l-arginine enables the nanomotor to have the ability to chemotaxis towards high concentrations of ROS/iNOS in tumors and be catalyzed to produce NO, achieving deep penetration into tumor tissue. Furthermore, the disulfide bond (SS) carried by the crosslinking agent used in the preparation of the nanomotor can respond to the high expression of reducing glutathione in the tumor microenvironment and undergo degradation, releasing a large amount of loaded drug DOX. Cell and animal disease model experiments confirme the good therapeutic effect of this drug-loaded nanomotor, providing new therapeutic concepts and strategies for the treatment of peritoneal metastatic cancer.

PH responsive and zwitterionic micelle for enhanced cellular uptake and antitumor performance

The side effects of small molecule chemotherapeutic drugs (SMCD) have brought great pain to the cancer patients. Many nanodrug carriers can relieve the shortcomings of SMCD, but they have complex synthesis processes and lack biodegradability. To overcome both problems, we synthesized a pH responsive biodegradable zwitterionic molecules (EK-D) by linking zwitterionic polypeptide (EK7) and dodecyl acrylate through a simple click reaction. Subsequently, doxorubicin (DOX) was physically encapsulated within the EK-D micelles to produce EK-D-DOX micelles, and polyethylene glycol monooleate (POO) employed as a comparative group for the preparation of POO-DOX micelles. The results show that EK-D-DOX micelles have good aqueous stability and anti-protein non-specific adsorption performance at pH 7.4, but EK-D-DOX micelles aggregate under the condition of pH = 5.5 due to the biodegradability of EK-D. The tumor cell uptake rate of EK-D-DOX micelles is higher than that of POO-DOX micelles and free DOX, which makes EK-D-DOX micelles the highest cytotoxic. Additionally, EK-D-DOX micelles release more DOX in a slightly acidic environment than at pH 7.4, and the release of DOX reaches a significant cumulative value of 75.20 % under pH conditions of 5.5. More importantly, EK-D-DOX micelles exhibit superior in vivo tumor inhibitory efficacy compared to free DOX, resulting in a remarkable tumor inhibition rate of 95.7 %. EK-D-DOX micelles not only have lower biological toxicity to normal tissues than free DOX, but also have a longer blood circulation time in mice. The method of EK-D-DOX micelles preparation represents a new method to prepare biodegradable zwitterionic nanodrug.

Fabrication of a Heptapeptide-Modified Poly(glycidyl Methac-Rylate) Nanosphere for Oriented Antibody Immobilization and Immunoassay

Oriented antibody immobilization has been widely employed in immunoassays and immunodiagnoses due to its efficacy in identifying target antigens. Herein, a heptapeptide ligand, HWRGWVC (HC7), was coupled to poly(glycidyl methacrylate) (PGMA) nanospheres (PGMA-HC7). The antibody immobilization behavior and antigen recognition performance were investigated and compared with those on PGMA nanospheres by nonspecific adsorption and covalent coupling via carbodiimide chemistry. The antibodies tested included bovine, rabbit, and human immunoglobulin G (IgG), while the antigens included horseradish peroxidase (HRP) and β-2-Microglobulin (β2-MG). The nanospheres were characterized using zeta potential and particle size analyzers, scanning electron microscopy, transmission electron microscopy, Fourier transform infrared spectroscopy, and reversed-phase chromatography, proving each synthesis step was succeeded. Isothermal titration calorimetry assay demonstrated the strong affinity interaction between IgG and PGMA-HC7. Notably, PGMA-HC7 achieved rapid and extremely high IgG adsorption capacity (~3 mg/mg) within 5 min via a specific recognition via HC7 without nonspecific interactions. Moreover, the activities of immobilized anti-HRP and anti-β2-MG antibodies obtained via affinity binding were 1.5-fold and 2-fold higher than those of their covalent coupling counterparts. Further, the oriented-immobilized anti-β2-MG antibody on PGMA-HC7 exhibited excellent performance in antigen recognition with a linear detection range of 0–5.3 μg/mL, proving its great potential in immunoassay applications.

An Effective Surface Passivation Assay for Single-Molecule Studies of Chromatin and Topoisomerase II.

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All-solid-state K+ sensing array based on Au@polystyrene nanocomposites.

All-solid-state ion selective electrodes (ASS-ISEs) areeasy to miniaturize and array, meeting the needs of home sensing devices. However, ASS-ISEs still faces challenges in accuracy and stability due to basic potential changes caused by non-specific adsorption of charged background compositions and the complex electrode preparation steps. To this end, our group successfully subtracted the background signal by integrating a self-calibrating channel in the sensing array and simplified the electrode preparation steps by preparing multi-functional PS-Au nanocomposites. However, the uniformity and gold content of PS-Au nanocomposites are difficult to control, so Au@PS nanocomposites are prepared as sensor materials in this paper to further reduce the differences between batches of electrodes. K+ Au@PS sensing array can be obtained by directly dropping Au@PS nanocomposites on the screen-printed carbon electrodes (SPCEs), which shows a near Nernstian behavior in the range 1.0 × 10-3M to 0.3M and good reproducibility in real sample testing. The detection results by K+ Au@PS sensing array for K+ in human morning urine agreed well with that tested by ICP-AES, which make the K+-ASS-ISE suitable for home health monitoring.

Ratiometric Electrochemical Detection of Interleukin-6 Using Electropolymerized Methylene Blue and a Multi-Walled Carbon-Nanotube-Modified Screen-Printed Carbon Electrode.

Herein, we report a ratio-based electrochemical biosensor for the detection of interleukin-6 (IL-6). We electropolymerized methylene blue (MB) on the surface of screen-printed carbon electrodes; introduced an internal reference signal probe; modified the carboxylate multi-walled carbon nanotubes on the electrode surface to increase the electrochemically active area; and finally linked the amino-modified IL-6 aptamer to the electrode surface through the Schiff base reaction, with bovine serum albumin (BSA) added to mask non-specific adsorption. After adding IL-6 to the samples, the signal of IMB remained almost unchanged, while the signal of I[Fe(CN)6]3-/4- decreased with increasing IL-6 concentration. Thus, a novel ratiometric electrochemical sensor with a linear range of 0.001~1000.0 ng/mL and a low detection limit of 0.54 pg/mL was successfully developed. The sensor had high repeatability, stability, sensitivity, and practicability. It provides a new method for constructing proportional electrochemical sensors and detecting IL-6.

Waveguide-Enhanced Nanoplasmonic Biosensor for Ultrasensitive and Rapid DNA Detection.

DNA is fundamental for storing and transmitting genetic information. Analyzing DNA or RNA base sequences enables the identification of genetic disorders, monitoring gene expression, and detecting pathogens. Traditional detection techniques like polymerase chain reaction (PCR) and next-generation sequencing (NGS) have limitations, including complexity, high cost, and the need for advanced computational skills. Therefore, there is a significant demand for enzyme-free and amplification-free strategies for rapid, low-cost, and sensitive DNA detection. DNA biosensors, especially those utilizing plasmonic nanomaterials, offer a promising solution. This study introduces a novel DNA-functionalized waveguide-enhanced nanoplasmonic optofluidic biosensor using a nanogold-linked sorbent assay for enzyme-free and amplification-free DNA detection. Integrating plasmonic gold nanoparticles (AuNPs) with a glass planar waveguide (WG) and a microfluidic channel, fabricated through cost-effective, vacuum-free methods, the biosensor achieves specific detection of complementary target DNA sequences. Utilizing a sandwich architecture, AuNPs labeled with detection DNA probes enhance sensitivity by altering evanescent wave distribution and inducing plasmon resonance modes. The biosensor demonstrated exceptional performance in DNA detection, achieving a limit of detection (LOD) of 33.1 fg/mL (4.36 fM) with a rapid response time of approximately 8 min. This ultrasensitive, rapid, and cost-effective biosensor exhibits minimal background nonspecific adsorption, making it highly suitable for clinical applications and early disease diagnosis. The innovative design and fabrication processes offer significant advantages for mass production, presenting a viable tool for precise disease diagnostics and improved clinical outcomes.