Immobilization Of Biomolecules Research Articles

Exploring various carbon nanomaterials-based electrodes modified with polymelamine for the reagentless electrochemical immunosensing of Claudin18.2

Claudin18.2 (CLDN18.2) is a tight junction protein often overexpressed in various solid tumors, including gastrointestinal and esophageal cancers, serving as a promising target and potential biomarker for tumor diagnosis, treatment assessment, and prognosis. Despite its significance, no biosensor has been reported to date for the detection of CLDN18.2. Here, we present the inaugural immunosensor for CLDN18.2. In this study, an amine-rich conducting polymer of polymelamine (PM) was electrografted onto different carbon nanomaterial-based screen-printed electrodes (SPEs), including carbon (C), graphene (Gr), graphene oxide (GO), carbon nanotube (CNT), and carbon nanofiber (CNF) via cyclic voltammetry. A comparative study was performed to explore the best material for the preparation of the PM-modified electrodes to be used as in-situ redox substrate for the immunosensor fabrication. The surface chemistry and structural features of pristine and PM-deposited electrodes were analyzed using Raman and scanning electron microscopy (SEM) techniques. Our results showed that the PM deposited on Gr and CNT/SPEs exhibited the most significant and stable redox behavior in PBS buffer. The terminal amine moieties on the PM-modified electrode surfaces were utilized for immobilizing anti-CLDN18.2 monoclonal antibodies via N-ethyl-N′-(3-(dimethylamino)propyl)carbodiimide/N-hydroxysuccinimide chemistry to construct the electrochemical immunosensor platform. Differential pulse voltammetry-based immunosensing of CLDN18.2 protein on BSA/anti-CLDN18.2/PM-Gr/SPE and BSA/anti-CLDN18.2/PM-CNT/SPE exhibited excellent selectivity against other proteins such as CD1, PDCD1, and ErBb2. The limits of detection of these two immunosensor platforms were calculated to be 7.9 pg/mL and 0.104 ng/mL for the CNT and Gr immunosensors, respectively. This study demonstrated that the PM-modified Gr and CNT electrodes offer promising platforms not only for the reagentless signaling but also for covalent immobilization of biomolecules. Moreover, these platforms offer excellent sensitivity and selectivity for the detection of CLDN18.2 due to its enhanced stable redox activity. The immunosensor demonstrated promising results for the sensitive detection of CLDN18.2 in biological samples, addressing the critical need for early gastric cancer diagnosis.

Nanostructured Metal Oxide-Based Electrochemical Biosensors in Medical Diagnosis.

Nanostructured metal oxides (NMOs) provide electrical properties such as high surface-to-volume ratio, reaction activity, and good adsorption strength. Furthermore, they serve as a conductive substrate for the immobilization of biomolecules, exhibiting notable biological activity. Capitalizing on these characteristics, they find utility in the development of various electrochemical biosensing devices, elevating the sensitivity and selectivity of such diagnostic platforms. In this review, different types of NMOs, including zinc oxide (ZnO), titanium dioxide (TiO2), iron (II, III) oxide (Fe3O4), nickel oxide (NiO), and copper oxide (CuO); their synthesis methods; and how they can be integrated into biosensors used for medical diagnosis are examined. It also includes a detailed table for the last 10 years covering the morphologies, analysis techniques, analytes, and analytical performances of electrochemical biosensors developed for medical diagnosis.

MOF-derived sandwich-structured dual Z-Scheme Co9S8@ZnIn2S4/CdSe hollow nanocages heterojunction: Target-induced ultrasensitive photoelectrochemical sensing of chlorpyrifos

Exploring efficient photoactive material presents an intriguing opportunity to enhance the analytical performance of photoelectrochemical (PEC) sensor in the environmental analysis. In this work, a sandwich-structured multi-interface Co9S8@ZnIn2S4/CdSe QDs dual Z-Scheme heterojunction, derived from metal-organic framework (MOF), was synthesized as a sensing platform for chlorpyrifos detection, by integrating with enzyme-induced in situ insoluble precipitates strategy. The meticulously designed Co9S8@ZnIn2S4/CdSe QDs exhibited enhanced charge separation efficiency and was proved to be a highly effective sensing platform for the immobilization of biomolecules, attributing to the intrinsic dual Z-Scheme heterojunction and the distinctive hollow structure. The proposed PEC sensing platform combined with enzyme-induced in situ precipitate signal amplification strategy achieved superior performance for sensing of chlorpyrifos (CPF), showing in wide linear range (1.0 pg mL−1–100 ng mL−1), with a limit of detection (0.6 pg mL−1), excellent selectivity, and stability. This work offers valuable insights for the design of novel advanced photoactive materials aimed at detecting environmental pollutants with low level concentration.

Reagent‐Free Covalent Immobilization of Biomolecules in a Microfluidic Organ‐On‐A‐Chip

AbstractMicrofluidic systems have become integral for lab‐on‐a‐chip and organ‐on‐a‐chip applications across numerous disciplines. These systems, typically fabricated using polydimethylsiloxane (PDMS) chips on glass substrates, lack the bioactivity required for such applications. To overcome this, biomolecules are immobilized using either oxygen (O2) plasma treatment or chemical reagents like amino silanes. However, O2 plasma treatments are unstable and cannot covalently immobilize biomolecules, while wet‐chemistry approaches are toxic, time‐consuming, and expensive. A novel microfluidic platform that combines two plasma surface treatments: Plasma‐activated coating (PAC) and atmospheric pressure plasma jet (APPJ), to enable reagent‐free covalent immobilization of biomolecules is described here. These surface treatments, unlike O2 plasma, covalently immobilized fibronectin on PDMS and glass, and significantly improved endothelial cell attachment and proliferation. By combining PAC and APPJ, a hybrid microfluidic platform with equivalent bond strength to standard O2 plasma devices, but with significantly enhanced endothelial cell growth in and artery‐on‐a‐chip model, is developed. This platform is also amenable to high‐shear applications such as coronary shear, with endothelial cells aligning with flow, as seen in human arteries. By providing reagent‐free covalent immobilization of biomolecules within a microfluidic system, this technology has the potential to radically improve organ‐on‐a‐chip development as well as lab‐on‐a‐chip systems, point‐of‐care diagnostics, and sensors.

High‐Resolution 3D Printing of Dual‐Curing Thiol‐Ene/Epoxy System for Fabrication of Microfluidic Devices for Bioassays

AbstractThe customized processing of polymer materials in the microscale by high‐resolution 3D printing provides an easy access to advanced applications in the fields of optics, microfluidics, tissue engineering, and life science. However, the 3D printing of enclosed structures in the scale of tens of microns such as closed microfluidic channels remains a challenge as channel structures often are clogged by residual cured resin. Dual‐curing systems based on off‐stoichiometric thiol‐ene and thiol‐ene/epoxy chemistry are well‐known for adhesive‐free bonding in the fabrication of cast or injection molded microfluidic devices. Herein, the first high‐resolution stereolithography 3D printing of a dual‐curing thiol‐ene/epoxy system in the microscale for the fabrication of customized microfluidic devices is presented. In the first curing step, by high‐resolution 3D printing open microfluidic structures are produced. Consecutively, the microchannels are sealed by adhesive‐free dry bonding upon thermal initiation, producing well‐controlled structures with channel sizes down to 80 µm. Before bonding, the intermediate material allows for tailored surface modification with biotin, which allows for consecutive immobilization of various biomolecules. A DNA‐bioassay with specific patterning is shown in the sealed chip. The presented work paves the way toward the fabrication of customized microfluidic devices for a large range of specific bioassays.

Graphene derivative-based ink advances inkjet printing technology for fabrication of electrochemical sensors and biosensors

The field of biosensing would significantly benefit from a disruptive technology enabling flexible manufacturing of uniform electrodes. Inkjet printing holds promise for this, although realizing full electrode manufacturing with this technology remains challenging. We introduce a nitrogen-doped carboxylated graphene ink (NGA-ink) compatible with commercially available printing technologies. The water-based and additive-free NGA-ink was utilized to produce fully inkjet-printed electrodes (IPEs), which demonstrated successful electrochemical detection of the important neurotransmitter dopamine. The cost-effectiveness of NGA-ink combined with a total cost per electrode of $0.10 renders it a practical solution for customized electrode manufacturing. Furthermore, the high carboxyl group content of NGA-ink (13 wt%) presents opportunities for biomolecule immobilization, paving the way for the development of advanced state-of-the-art biosensors. This study highlights the potential of NGA inkjet-printed electrodes in revolutionizing sensor technology, offering an affordable, scalable alternative to conventional electrochemical systems.

Dielectrically-Modulated GANFET Biosensor for Label-Free Detection of DNA and Avian Influenza Virus: Proposal and Modeling

This paper introduces a novel device called the Gate All Around Engineered Gallium Nitride Field Effect Transistor (GAAE-GANFET), designed specifically for label-free biosensing applications. This innovative gate-all-around engineering in GANFET integrates various device engineering techniques, such as channel engineering, gate engineering, and oxide engineering, to enhance biosensing performance. The channel engineering techniques refer to the use of a gallium nitride channel with a step-graded doping profile, divided into three distinct regions. In contrast, the gate engineering technique refers to the cylindrical split-gate-underlap architecture. The oxide engineering technique involves stacking Al2O3 and HfO2. Moreover, this biosensor incorporates two-sided gate underlap cavities that facilitate the immobilization of biomolecules. These open cavities not only provide structural stability but also simplify the fabrication process to a significant extent. The viability of this biosensor as a label-free biosensor has been evaluated using an antigen and an antibody from the Avian Influenza virus and DNA as the target biomolecules. The proposed analytical model and TCAD simulation results are in excellent agreement, demonstrating the reliability of the proposed device. Additionally, the biosensor’s sensitivity, which depends on cavity length, doping concentration, gate metal work function, and temperature variation, has been thoroughly explored. The gate-all-around structure, along with the integration of tri-step graded doping, GaN as the channel material, gate oxide stacking, and dual open cavity structure in the proposed biosensor, leads to significantly improved biosensing capabilities.

Sandwich-type electrochemical aptasensor based on hemin-graphite oxide as a signal label and rGO/MWCNTs/chitosan/carbon quantum dot modified electrode for sensitive detection of Acinetobacter baumannii bacteria

Acinetobacter baumannii (A. baumannii) is a pathogenic bacterium that causes severe infections and its rapid and reliable diagnosis is essential for effective control and treatment. In this study, we present an electrochemical aptasensor based on a signal amplification strategy for the detection of A. baumannii, the high specificity and affinity of the aptamer for the target make it favorable for signal amplification. This allows for a highly sensitive and selective detection of the target. The aptasensor is based on a carbon screen-printed electrode (CSPE) that has been modified with a nanocomposite consisting of multi-walled carbon nanotubes (MWCNTs), reduced graphene oxide (rGO), chitosan (CS), and a synthesized carbon quantum dot (CQD) from CS. Additionally, the self-assembled aptamers were immobilized on hemin-graphite oxide (H-GO) as a signal probe.The composition of the nanocomposite (rGO-MWCNT/CS/CQD) provides high conductivity and stability, facilitating the efficient capture of A. baumannii onto the surface of the aptasensor. Also, aptamer immobilized on Hemin-graphite oxide (H-GO/Aptamer) was utilized as an electrochemical signal reporter probe by H reduction. This approach improved the detection sensitivity and the aptamer surface density for detecting A. baumannii. Furthermore, under optimized experimental conditions, the aptasensor was demonstrated to be capable of detecting A. baumannii with a linear range of (10 – 1 × 107 Colony-forming unit (CFU)/mL) and a limit of detection (LOD) of 1 CFU/mL (σ = 3). One of the key features of this aptasensor is its ability to distinguish between live and dead bacteria cells, which is very important and critical for clinical applications. In addition, we have successfully detected A. baumannii bacteria in healthy human serum and skim milk powder samples provided using the prepared electrochemical aptasensor.The functional groups present in the synthetic CQD, rGO-MWCNT, and chitosan facilitate biomolecule immobilization and enhance stability and activity. The fast electron-transfer kinetics and high conductivity of these materials contribute to improved sensitivity and selectivity. Furthermore, The H-GO/Aptamer composite's large surface area increases the number of immobilized secondary aptamers and enables a more stable structure. This large surface area also facilitates more H loading, leading to signal amplification.

Ultrasensitive and label-free electrochemical immunosensor for the detection of the ovarian cancer biomarker CA125 based on CuCo-ONSs@AuNPs nanocomposites

Cancer antigen 125 (CA125) is pivotal as a tumor marker in early ovarian cancer prevention and diagnosis. In this work, we introduced an ultrasensitive label-free electrochemical immunosensor tailored for CA125 detection, leveraging nanogold-functionalized copper-cobalt oxide nanosheets (CuCo-ONSs@AuNPs) as nanocomposites. For the inaugural application, copper-cobalt oxide nanosheets delivered the requisite DPV electrochemical response for the immunosensors. Their large specific surface area and commendable electrical conductivity amplify electron transfer and enable significant gold nanoparticle loading. Concurrently, AuNPs offer a plethora of active sites, facilitating easy immobilization of biomolecules via the bond between amino groups and AuNPs. We employed scanning electron microscopy, transmission electron microscopy, and x-ray photoelectron spectroscopy to characterize the nanomaterials' surface morphology and elemental composition. The electrochemical sensor response signals were ascertained using differential pulse voltammetry. Under optimal conditions, the immunosensor exhibited a linear detection range from 1×10−7 U/mL to 1×10−3 U/mL and a detection limit of 3.9×10−8 U/mL (S/N=3). The proposed label-free electrochemical immunosensor furnishes a straightforward, dependable, and sensitive approach for CA125 quantification and stands as a promising method for clinical detection of other tumor markers.

Itaconic Acid Cross-Linked Biomolecule Immobilization Approach on Amine-Functionalized Silica Nanoparticles for Highly Sensitive Enzyme-Linked Immunosorbent Assay (ELISA).

Biomolecule immobilization on nanomaterials is attractive for biosensors since it enables the capture of a higher concentration of bioreceptor units while also serving as a transduction element. The technique could enhance the accuracy, specificity, and sensitivity of the analytical measurements of biomolecules. However, it was found that the limitation in chemically binding biomolecules on nanoparticle surfaces could only cross-link between the C-terminal and N-terminal. Here, we report the facile one-step synthesis of amine-functionalized silica nanoparticles (AFSNPs). (3-Aminopropyl)triethoxysilane was used as a precursor to modify the functional surface of nanoparticles via the Stöber process. The biomolecules were immobilized to the AFSNPs through itaconic acid, a novel cross-linker that binds between the N-terminal and N-terminal and potentially improves proteins and nucleic acid immobilization onto the nanoparticle surface. The newly developed immobilization approach on AFSNPs for biomolecular detection enhanced the efficiency of ELISA, resulting in increased sensitivity. It might also be easily used to identify different pathogens for clinical diagnostics.

Polymer brush construction with interface-mediated RAFT polymerization technique for glutathione determination

Reactive polymeric thin films designed on solid surfaces have widespread applications in many fields involving the immobilization of biomolecules. Efficient and specific designs for biomolecule immobilization on polymer-modified solid surfaces are central to biosensor technologies such as protein and gene chips. Research areas related to the functionalization of surfaces include polymer brushes whose functionality is increased by the Diels-Alder (DA) reaction, which have been made usable as rewritable platforms with their thermal transformation properties and have found a lot of use, especially in biomolecule determination studies. Glutathione (GSH), one of the biomolecules, is found in in very high intracellular concentrations. It is the most important antioxidant molecule in the environment and its determination is a good indicator for us to obtain information about the status of the antioxidant defense mechanism in various diseases. When studies in were examined in detail, no study was found on the synthesis of polymer brushes with thiol functional groups covalently bonded to the silicon surface using the interfacial-mediated RAFT polymerization technique and their use in GSH adsorption. Therefore, in this study, to fill this gap in the literature, polymer brushes with thiol (SH) functional groups to be used in GSH adsorption were synthesized. Poly(furfurylmethacrylate) [poly(FMA)] brushes, formed by polymer chains covalently bonded to the silicon surface, were synthesized by interface-mediated RAFT polymerization. Then, by maleimide conjugation using hexylamine, polymeric brushes with thiol functional groups covalently bonded to the silicon surface which can be used in GSH adsorption were obtained for determination purposes. It is thought that the prepared polymeric brushes will find a wide application area for the determination of many new molecules, especially the determination of glutathione and will contribute to the literature.

Robust Gold Electrode Fabrication in a Microfluidic System

Electrode fabrication in microfluidic devices is often avoided due to the smaller footprint of the platform, associated complexity in the process, and absence of a well-established method. In this article a glass microchip with chemically modified and vapor deposited gold internal electrode was fabricated and used for biomolecule immobilization and electrochemical applications. The silanol (-OH) on glass surface of the microdevice was modified with (3-mercaptopropyl)trimethoxysilane prior to gold deposition to achieve a strong attachment of the metal film using the gold-thiol binding. The practical utility of the microdevice with gold surface was demonstrated by performing quantitative enzyme linked immunosorbent assay (ELISA) of human interferon gamma (IFN-γ) antigen, yielding a limit of detection (LOD) of 0.15 pg/mL compared to 4 pg/mL in the microplate assay. Additionally, the functionality of the internal electrode was established by performing cyclic voltammetry (CV) and linear sweep voltammetry (LSV) of potassium ferricyanide (III) in potassium chloride (KCl) solution using the proposed gold patterned electrode in the microchip, offering limits of detection of 0.14 and 0.11 mM for CV and LSV respectively. Both the ELISA and electrochemical assays were validated by generating calibration curves and using a control sample for the electroanalytical sensing applications. The measurements yielded experimental results that agreed with control concentration with 7% relative standard deviation (RSD). The promising reproducibility of the electrode in fabrication with no/minimum failure rate and its outstanding performance in immunosensor applications as well as an electron exchange surface makes our technique of gold electrode fabrication in microchips a state of the art.

Development of gold nanospikes-modified quartz crystal microbalance biosensor for prostate specific antigen detection

Prostate-specific antigen (PSA) screening is vital for the early detection and assessment of prostate cancer treatment efficacy. Integrating nanotechnology into biosensor development allows for targeted biomarker detection. In this study, we present the fabrication of gold nanospikes (AuNS) structures on a quartz crystal microbalance (QCM) using the pulse current electrodeposition (PCD) technique. By varying the duty cycles of PCD, we achieved the desired nanostructure. The characteristics of the AuNS-modified QCM electrode were thoroughly investigated using various micro-structural characterization techniques such as x-ray diffraction (XRD), field emission scanning electron microscope (FESEM), atomic force microscope (AFM), X-ray photoelectron spectroscopy (XPS) and cyclic voltammetry (CV). Prior to biomolecule immobilization, each chemical treatment was verified through contact angle and FT-IR analyses. The performance of the developed QCM biosensor was evaluated using PSA at various concentrations. The results revealed that the frequency variation of the QCM biosensor was directly proportional to the PSA concentration. Notably, the AuNS-modified QCM biosensor exhibited the highest frequency shift at a 20 % duty cycle, demonstrating a linear detection range of 0.1–100 ng mL−1 and a limit of detection (LOD) of 24 pg mL−1. The developed QCM biosensor significantly enhances detection efficiency by increasing the number of binding sites for biomolecules through an enlarged surface area. The demonstrated sensitivity of the AuNS-modified QCM biosensor indicates its potential to address the increasing demand for early diagnosis and screening in healthcare systems. This innovative biosensor technology holds promise for improving prostate cancer detection and has broader implications for the development of highly sensitive biosensors for various biomarkers.

Review—Potential of Tunneling Magnetoresistance Coupled to Iron Oxide Nanoparticles as a Novel Transducer for Biosensors-on-Chip

Biosensors-on-chip (BoC), compact and affordable public diagnostic devices, are vital for preventing health crises caused by viral and bacterial mutations, climate change, and poor diets. Clinical, remote, and field use are possible with these devices. BoC is used in food safety, environmental monitoring, and medical diagnosis. The coupling of tunneling magnetoresistance (TMR) sensing elements in chip form with surface functionalized iron oxide nanoparticles (IONPs) as a biomarker, known as TMR/ IONPs, allows BoC devices to be made. The functional framework of BoC based on TMR/ IONPs, the instrument system, and biomolecule immobilization will be covered in this review. This review aims to overview the recent research on a biosensor using TMR technology with IONPs biomarkers and discuss its future advances in point-of-care diagnostics. TMR sensors have revolutionized low-magnetic field sensing technologies, yet biosensing faces challenges. However, it’s important to recognize that TMR sensors exclusively utilize out-of-plane connections. that creates a larger separation between biomolecules and the sensing layer, leading to a decrease in the sensitivity of biomolecules. Large magnetization of dispersed IONPs to generate sufficient stray-field, compact and inexpensive instrumentation to sense the low voltage yielded by the TMR/IONPs system, and high-selectivity bio-analyte immobilization to the surface of IONPs to increase sensor sensitivity are the notable issues to address this problem. The utilization of the green synthesized method in producing IONPs for magnetic labeling can enhance the sensitivity of a TMR-based biosensor. This is achieved through the formation of IONPs with a capping agent on its surface that inhibits the aggregation and facilitates the immobilization of the biomolecule target. Additionally, improving device performance requires creating high-TMR materials. Despite challenges, research and technological advances hold great promise for TMR/GS-IONP bio-applications.

4-(Triethoxysilyl)butanoic acid as a self-assembled monolayer for surface modification of titanium dioxide.

In this study, the carboxy silane 4-(triethoxysilyl)butanoic acid (TESBA) was used to modify titanium dioxide (TiO2) to create a self-assembled monolayer (SAM) and then directionally immobilize a capture antibody using protein A. We selected the amino silane (3-aminopropyl)triethoxysilane (APTES) to perform a comparative analysis with TESBA, and employed glutaraldehyde (GA) as the control. The modification and detection effects and the limit of detection (LOD) were evaluated by detecting human immunoglobulin G (IgG). The average normalized sensitivity of the dual-grating coupler waveguide biosensor was 49.63 ± 0.27 RIU-1 and the optimum resolution was 1.30 × 10-6 RIU. When the SAM was prepared using TESBA and APTES followed by GA, the LOD was 4.59 × 10-7 g mL-1 and 5.29 × 10-7 g mL-1, respectively. We analyzed the modification and detection effects by the t-test and concluded that the differences in the modification effects using TESBA and APTES followed by GA were significant and the differences in the detection effects using TESBA and APTES followed by GA were insignificant. The use of TESBA as the SAM led to the modification effect being superior to that obtained using APTES followed by GA. The detection effect using TESBA was as outstanding as that using APTES followed by GA. Our findings demonstrate the feasibility and effectiveness of using TESBA as the SAM to carboxylate the surface of TiO2, thereby enabling immobilization of biomolecules for human IgG detection.

Nano-modified screen-printed electrode-based electrochemical immunosensors for oral cancer biomarker detection in undiluted human serum and saliva samples.

This proposed work reports the development of in-house made conductive ink-based screen-printed electrodes (SPEs) for label-free detection of oral cancer biomarkers. Carbon ink synthesis includes graphite powder, gum arabic, and water. The selectivity test of the fabricated SPE involves immobilizing antibodies specific to biomarkers and challenges with redox-active interference, other serum molecules, and non-target biomarkers. Three different biomarkers, cytokeratin-19 fragment (CYFRA 21-1), interleukin 8 (IL-8), and tumor protein p53 (TP-53), act as target entities for the detection of oral cancer in patients' samples (serum, N = 28, and saliva, N = 16) at an early stage. The standard technique enzyme-linked immunosorbent assay (ELISA) was employed to estimate the concentration of the biomarkers in serum and saliva samples. SPEs contain amine (-NH2) functional groups involved in covalent bonding with the carboxyl (-COOH) groups of antibody molecules. These immunosensors exhibited remarkably lower detection limits of 829.5 pg mL-1, 0.543 pg mL-1, and 1.165 pg mL-1, and excellent sensitivity of 0.935 μA mL pg-1 cm-1, 0.039 μA mL pg-1 cm-1, and 0.008 μA mL pg-1 cm-1 for CYFRA 21-1, IL-8, and TP-53 biomarkers, respectively. This sensing platform does not require any functionalization for biomolecule immobilization. Thus, it is a cost-effective, disposable, flexible, miniaturized, and sensitive strip to detect oral cancer biomarkers.

Functionalization of Polydimethylsiloxane with Diazirine-Based Linkers for Covalent Protein Immobilization.

Biomolecule attachment to solid supports is critical for biomedical devices, such as biosensors and implants. Polydimethylsiloxane (PDMS) is commonly used for these applications due to its advantageous properties. To enhance the biomolecule immobilization on PDMS, a novel technique is demonstrated using newly synthesized diazirine molecules for the surface modification of PDMS. This nondestructive process involves a reaction between diazirine molecules and PDMS through C-H insertion with thermal or ultraviolet activation. The success of the PDMS modification is confirmed by various surface characterization techniques. Bovine serum albumin (BSA) and immunoglobulin G (IgG) are strongly attached to the modified PDMS surfaces, and the amount of protein is quantified using iodine-125 radiolabeling. The results demonstrate that PDMS is rapidly functionalized, and the stability of the immobilized proteins is significantly improved with multiple types of diazirine molecules and activation methods. Confocal microscopy provides three-dimensional images of the distribution of immobilized IgG on the surfaces and the penetration of diazirine-based linkers through the PDMS substrate during the coating process. Overall, this study presents a promising new approach for functionalizing PDMS surfaces to enhance biomolecule immobilization, and its potential applications can extend to multimaterial modifications for various diagnostic and medical applications such as microfluidic devices and immunoassays with relevant bioactive proteins.

Electrochemical Sensing Based on Nanofibers Modified Electrodes for Application in Diagnostic, Food and Waste Water Samples

AbstractElectrochemical sensors and biosensors are today important analytical and monitoring tools in various fields, from agriculture and the food industry to environmental and biomedical/pharmaceutical applications. In particular, the integration of nanotechnology with electrochemical sensors and biosensors to develop a new generation of sensor platforms has made enormous progress in recent years. The outstanding properties of one‐dimensional (1D) nanofibers (NFs), such as high porosity, superior mechanical properties and high specific surface area have made them attractive electrocatalysts, support materials for the immobilization of biomolecules as well as mimetic materials for sensing and biosensing applications. Moreover, the possibility of fabricating multifunctional composites based on NFs increases (bio)sensing capabilities through synergistic effects and additive properties. This review describes the progress made over the last decade in the use of multifunctional NFs‐based composites as modified electrodes for the sensing of various analytes in biomedical, food, and wastewater treatment applications. The aim of this review is to provide a comprehensive overview and a guide for researchers from different disciplines to fabricate and improve their selective NFs‐based (bio)sensor platforms for the detection of desired analytes or multi‐analytes.

Multifunctional Poly(2-ethyl-2-oxazoline) Copolymers Containing Dithiolane and Pentafluorophenyl Esters as Effective Reactive Linkers for Gold Surface Coatings.

Surface functionalization with biological macromolecules is an important task for the development of sensor materials, whereby the interaction with other biological materials should be suppressed. In this work, we developed a novel multifunctional poly(2-ethyl-2-oxazoline)-dithiolane conjugate as a versatile linker for gold surface immobilization of amine-containing biomolecules, containing poly(2-ethyl-2-oxazoline) as antifouling polymer, dithiolane for surface immobilization, and activated esters for protein conjugation. First, a well-defined carboxylic acid containing copoly(2-ethyl-2-oxazoline) was synthesized by cationic ring-opening copolymerization of 2-ethyl-2-oxazoline with a methyl ester-containing 2-oxazoline monomer, followed by postpolymerization modifications. The side-chain carboxylic groups were then converted to amine-reactive pentafluorophenyl (PFP) ester groups. Part of the PFP groups was used for the attachment of the dithiolane moiety, which can efficiently bind to gold surfaces. The final copolymer contained 1.4 mol% of dithiolane groups and 4.5 mol% of PFP groups. The copolymer structure was confirmed by several analytical techniques, including NMR spectroscopy and size-exclusion chromatography. The kinetics of the PFP ester aminolysis and hydrolysis demonstrated significantly faster amidation compared to hydrolysis, which is essential for subsequent protein conjugation. Successful coating of gold surfaces with the polymer was confirmed by spectroscopic ellipsometry, showing a polymer brush thickness of 4.77 nm. Subsequent modification of the coated surfaces was achieved using bovine serum albumin as a model protein. This study introduces a novel reactive polymer linker for gold surface functionalization and offers a versatile polymer platform for various applications including biosensing and surface functionalization.

Using dynamic biomaterials to study the temporal role of bioactive peptides during osteogenesis

The extracellular matrix is a highly dynamic environment, and the precise temporal presentation of biochemical signals is critical for regulating cell behavior during development, healing, and disease progression. To mimic this behavior, we developed a modular DNA-based hydrogel platform to enable independent and reversible control over the immobilization of multiple biomolecules during in vitro cell culture. We combined reversible DNA handles with a norbornene-modified hyaluronic acid hydrogel to orthogonally add and remove multiple biomolecule-DNA conjugates at user-defined timepoints. We demonstrated that the persistent presentation of the cell adhesion peptide RGD was required to maintain cell spreading on hyaluronic acid hydrogels. Further, we discovered the delayed presentation of osteogenic growth peptide (OGP) increased alkaline phosphatase activity compared to other temporal variations. This finding is critically important when considering the design of OGP delivery approaches for bone repair. More broadly, this platform provides a unique approach to tease apart the temporal role of multiple biomolecules during development, regeneration, and disease progression.