Cysteamine Self-assembled Monolayer Research Articles

Screen-printed carbon electrode functionalized with AuNPs-cysteamine self-assembled monolayers for enzymatic uric acid detection in non-invasive samples.

Uric acid (UA) is a relevant biomarker that at abnormal levels could provide information for the timely diagnosis of chronic-degenerative diseases, such as Diabetes mellitus, cardiovascular deficiencies or gut. This work presents a simple surface functionalization of screen-printed carbon electrodes (SPCE) with cysteamine self-assembled monolayers (SAMs) assembled over electrodeposited gold nanoparticles (AuNPs). The modification allowed the immobilization of uricase enzyme, preserving its biocatalytic activity and resulting in sensitive and selective UA detection. The developed biosensor device exhibited a linear detection range from 100 μM – 1000 μM, a sensitivity of 6.622 nA/μM and a limit of detection (LOD) of 4.6 μM with selectivity to UA molecules over common interfering analytes. When evaluated in urine samples, the analytical capabilities of the PTSPCE/AuNPS/SAMs/Uox biosensor remained, achieving an average recovery rate of 126.91%. The obtained analytical parameters proved to be competitive when utilizing non-invasive fluids, suggesting the possibility of conducting detection assays with potential clinical applications using an implemented electrochemical biosensor through a simple, flexible, and reproducible methodology.

Label-free electrochemical immunosensor for Ochratoxin a detection in coffee samples

Ochratoxin A (OTA) is a nephrotoxic and carcinogenic mycotoxin frequently found in coffee, which directly impacts human health and the economy of many countries. For this reason, there has been a growing need for simple and sensitive tools for the on-site detection of this mycotoxin. In this study, we developed a label-free impedimetric immunosensor to detect OTA. The biosensor was built on a thin-film gold electrode evaporated on glass substrtes, modified with a self-assembled cysteamine monolayer and anti-OTA antibodies. Atomic force microscopy and Microspectroscopy RAMAN confirmed the successful functionalization of the electrodes. The biosensor performance was evaluated by electrochemical impedance spectroscopy and the measurements indicated a linear relationship between the change in the impedance values and the OTA concentration in the range from 0.5 to 100 ng mL−1 with a limit of detection of 0.15 ng mL−1. The biosensor was highly selective and did not suffer matrix interference when analyzed in coffee samples. Furthermore, considering the small sample volumes, the short time required for analysis, and the possibility of miniaturization, the developed biosensor represents a promising analytical device for on-site coffee quality analyses.

Self-Assembled Monolayers on Screen-Printed Gold Electrodes for Uricase Enzyme Covalent Immobilization

This work reports the development of an electrochemical biosensor for uric acid (UA), based on covalent immobilization of uricase (Uox) on cysteamine self-assembled monolayers (SAM). The monolayers were prepared over screen-printed gold electrode (AuSPE) modified with Au nanostructures for covalent linkage through EDC-NHS. The modified AuSPE was characterized by Scanning Electron Microscope (SEM) and Cyclic Voltammetry (CV). The results verified the cysteamine SAM formation, and the improved electrochemical performance of the working surface. The AuSPE/SAM/Uox biosensor presented a linear range of 0.84 – 16.81 mg/dL (50-1000 µM), a sensibility of 9.1423 µA/(mg/dL)cm2, and a limit of detection (LOD) of 0.0700 mg/dL. The biosensor did not present a response due to ascorbic acid (AA), verifying the selectivity of the assembly only to UA oxidation. The AuSPE/SAM/Uox showed a promising performance as a reliable, simple and fast detection tool for UA.

Development of a quartz crystal microbalance-based immunosensor for the early detection of mesothelin in cancer

A robust real-time flow injection assay using Quartz Crystal Microbalance (QCM) biosensor has been developed for the detection of mesothelin, an antigen expressed on various malignant tumors including mesothelioma and ovarian cancers. A QCM sensor chip functionalized with self-assembled monolayer of cysteamine used to fabricate a sensitive immunosensor. Mesothelin specific antibody was immobilized on cysteamine modified gold surface of quartz crystal using N-ethyl-N’-(3-dimethylaminopropyl) carbodiimide and N-hydroxy succinimide coupling. Further, ethanolamine was used as a blocking reagent to prevent the non-specific adsorption on antibody functionalized gold surface. Various concentrations of mesothelin was tested and the resonant frequency variation of the crystal was observed by a quartz crystal microbalance until a stable response is obtained. A good correlation between the frequency changes and concentration of mesothelin tested was observed and the QCM biosensor detects mesothelin in the linear range of 100pg/mLto 50 ng/mL. Thus, QCM assay could be a promising technique for early diagnosis of mesothelioma, ovarian and pancreatic adenocarcinoma.

Electrochemical DNA Biosensor Based on Immobilization of a Non-Modified ssDNA Using Phosphoramidate-Bonding Strategy and Pencil Graphite Electrode Modified with AuNPs/CB and Self-Assembled Cysteamine Monolayer

The present paper describes an alternative approach to the traditionally used covalent immobilization methods that require cost-intensive and complicated chemistry modification of a single-stranded DNA (ssDNA) capture probe. The low-cost pencil graphite electrode (PGE) modified with carbon black (CB) and gold nanoparticles (AuNPs) was used as an electrochemical platform and the non-modified ssDNA was immobilized on a self-assembled cysteamine modified AuNPs/CB-PGE through a phosphoramidate bond between the 5'-terminal phosphate group of ssDNA and the primary amine group of cysteamine. The microRNA-21 was used as a target model in the fabrication of this electrochemical DNA biosensor and the hybridization process with the complementary probe was monitored by differential pulse voltammetry using methylene blue (MB) as an electrochemical hybridization indicator. The decreased reduction peak current of MB shows a good linear correlation with the increased concentration of microRNA-21 target sequences because the MB signal is determined by the amount of exposed guanine bases. The linear range of the fabricated DNA biosensor was from 1.0 × 10-8 to 5.0 × 10-7 M with a detection limit of 1.0 × 10-9 M. These results show that the covalent immobilization of a non-modified ssDNA capture probe through a phosphoramidate-bonding strategy could serve as a cost-effective and versatile approach for the fabrication of DNA biosensors related to a wide range of applications that cover the fields of medical diagnostic and environmental monitoring. The fabricated electrochemical DNA biosensor was used to analyze microRNA-21 in a (spiked) human serum sample and it showed satisfactory and encouraging results as an electrochemical DNA biosensor platform.

Organic Anion Detection with Functionalized SERS Substrates via Coupled Electrokinetic Preconcentration, Analyte Capture, and Charge Transfer.

Detecting ultralow concentrations of anionic analytes in solution by surface-enhanced Raman spectroscopy (SERS) remains challenging due to their low affinity for SERS substrates. Two strategies were examined to enable in situ, liquid phase detection using 5(6)-carboxyfluorescein (5(6)-FAM) as a model analyte: functionalization of a gold nanopillar substrate with cationic cysteamine self-assembled monolayer (CA-SAM) and electrokinetic preconcentration (EP-SERS) with potentials ranging from 0 to +500 mV. The CA-SAM did not enable detection without an applied field, likely due to insufficient accumulation of 5(6)-FAM on the substrate surface limited by passive diffusion. 5(6)-FAM could only be reliably detected with an applied electric field with the charged molecules driven by electroconvection to the substrate surface and the SERS intensity following the Langmuir adsorption model. The obtained limits of detection (LODs) with an applied field were 97.5 and 6.4 nM on bare and CA-SAM substrates, respectively. For the CA-SAM substrates, both the ligand and analyte displayed an ∼15-fold signal enhancement with an applied field, revealing an additional enhancement due to charge-transfer resonance taking place between the metal and 5(6)-FAM that improved the LOD by an order of magnitude.

Electrochemical Immunosensor for the Quantification of S100B at Clinically Relevant Levels Using a Cysteamine Modified Surface.

Neuronal damage secondary to traumatic brain injury (TBI) is a rapidly evolving condition, which requires therapeutic decisions based on the timely identification of clinical deterioration. Changes in S100B biomarker levels are associated with TBI severity and patient outcome. The S100B quantification is often difficult since standard immunoassays are time-consuming, costly, and require extensive expertise. A zero-length cross-linking approach on a cysteamine self-assembled monolayer (SAM) was performed to immobilize anti-S100B monoclonal antibodies onto both planar (AuEs) and interdigitated (AuIDEs) gold electrodes via carbonyl-bond. Surface characterization was performed by atomic force microscopy (AFM) and specular-reflectance FTIR for each functionalization step. Biosensor response was studied using the change in charge-transfer resistance (Rct) from electrochemical impedance spectroscopy (EIS) in potassium ferrocyanide, with [S100B] ranging 10–1000 pg/mL. A single-frequency analysis for capacitances was also performed in AuIDEs. Full factorial designs were applied to assess biosensor sensitivity, specificity, and limit-of-detection (LOD). Higher Rct values were found with increased S100B concentration in both platforms. LODs were 18 pg/mL(AuES) and 6 pg/mL(AuIDEs). AuIDEs provide a simpler manufacturing protocol, with reduced fabrication time and possibly costs, simpler electrochemical response analysis, and could be used for single-frequency analysis for monitoring capacitance changes related to S100B levels.

Voltammetry and Single‐Molecule In Situ Scanning Tunnelling Microscopy of the Redox Metalloenzyme Human Sulfite Oxidase

AbstractHuman sulfite oxidase (hSO) is a homodimeric two‐domain enzyme central in the biological sulfur cycle. A pyranopterin molybdenum cofactor (Moco) is the catalytic site and a heme b5 group located in the N‐terminal domain. The two domains are connected by a flexible linker region. Electrons produced at the Moco in sulfite oxidation, are relayed via heme b5 to electron acceptors or an electrode surface. Inter‐domain conformational changes between an open and a closed enzyme conformation, allowing “gated” electron transfer has been suggested. We first recorded cyclic voltammetry (CV) of hSO on single‐crystal Au(111)‐electrode surfaces modified by self‐assembled monolayers (SAMs) both of a short rigid thiol, cysteamine and of a longer structurally flexible thiol, ω‐amino‐octanethiol (AOT). hSO on cysteamine SAMs displays a well‐defined pair of voltammetric peaks around −0.207 V vs. SCE in the absence of sulfite substrate, but no electrocatalysis. hSO on AOT SAMs displays well‐defined electrocatalysis, but only “fair” quality voltammetry in the absence of sulfite. We recorded next in situ scanning tunnelling spectroscopy (STS) of hSO on AOT modified Au(111)‐electrodes, disclosing, a 2–5 % surface coverage of strong molecular scale contrasts, assigned to single hSO molecules, notably with no contrast difference in the absence and presence of sulfite. In situ STS corroborated this observation with a sigmoidal tunnelling current/overpotential correlation.

A simple immunosensor for thyroid stimulating hormone

Determination of thyroid-stimulating hormone (TSH) level in serum or plasma is defined as a sensitive method for the diagnosis of hyperthyroidism and hypothyroidism and also in many diseases thought to be related to TSH levels. In this study, a novel simple impedimetric immunosensor based on polyamidoamine dendrimer was developed. Anti TSH antibody was immobilized on the gold electrode by using cysteamine self-assembled monolayer strategy. In constructing the immunosensor, a polyamidoamine dendrimer was used to increase the surface area in which Antı-TSH was immobilized and glutaraldehyde was used as a cross-linker. After each immobilization step, the electrode surface was monitored by electrochemical impedance spectroscopy, cyclic voltammetry, scanning electron microscopy and energy-dispersive X-ray spectroscopy techniques and optimization studies were performed. The reproducibility, repeatability, linearity and sensitivity of the immunosensor were examined. Also, the interference experiments for glucose, salts and proteins in serum were performed. The limit of detection and limit of quantification values of the proposed immunosensor were 0.026 mIUL−1 and 0.086 mIUL−1, respectively and it was able to detect the amount of TSH within a linear range of 0.1–0.6 mIUL−1.

Homosistein tayini amaçlı QCM-temelli biyosensör

High plasma homocysteine levels can cause an increased risk of cardiovascular, cerebrovascular, and peripheral arterial diseases. Besides this, Alzheimer’s disease and other dementias, osteoporosis, diabetes and renal disease due to folate and B-vitamin deficiency, various drugs or pre-existing atherosclerotic diseases may be the result of high homocysteine levels. The presented research work aimedto perform the detection of Homocysteine (Hcy) by using Quartz Crystal Microbalance (QCM) biosensor. The temperature controlled QCM system was a home-made designed and constructed equipment which can use silver electrod quartz crystal. The modification of silver electrod quartz crystals surfaces was achieved by the surface cleaning process with sodium hydroxide, acetone and methanol in a consecutive manner. Then self-assembled monolayer of cysteamine and chemical coupling of glutaraldehyde (GA) to free end of monolayer was achieved to create the new functional surface in order to complete the formation of spacer arm/ligand. Homocysteine specific recognizing ligand, anti-Homocysteine antibody was immobilized to glutaraldehyde coupled surfaces. The change in resonance frequency values were measured for each modification step. The optimization of dilution ratio of the antibody solution was performed to modified surfaces. The least dilution ratio of antibody, 1/10000 v/v, was determined as optimum antibody ratio. The detection of homocystein was analysed at a detection limit of 0.1 µM and the linear ranges of calibration curves were estimated as 0.1-2.0 µM and 10-50 µM. Homocysteine values indicated good linearities (R2=0.9813 and 0.9875, respectively). The relative standart deviation (RSD %) for precision was calculated as less than 10%. In conclusion, it was found that the detection of homocysteine can be done both in nano- and micro-molar concentration levels. Additionally, designed biosensor showed desired stability and reproducibility. Finally, a new method different from the present methods for the use in the analysis of Hcy was proposed and developed which detects homocysteine by designed QCM technique with a rapid, cheeper and less pretreatment processes. Additionally, homocysteine detection was performed in nano- and micro- molar concentration values.

Nanoporous Thin Films Formed from Photocleavable Diblock Copolymers on Gold Substrates Modified with Thiolate Self-Assembled Monolayers.

Nanoporous thin films formed on electrodes are considered functional elements of electrochemical sensing systems, thus motivating methods for their development. We report a preparative strategy detailing the effects of surface modification of gold substrates with thiolate self-assembled monolayers (SAMs) on the properties of nanoporous thin films derived from polystyrene-block-poly(ethylene oxide) having a photocleavable o-nitrobenzyl ester junction (PS-hν-PEO). Two PS-hν-PEO having similar PEO volume fractions (≈0.2) but different molecular weights (10 and 23 kg/mol) were used to prepare films (30-100 nm thick) spin-cast on gold substrates unmodified and modified with cysteamine, thioctic acid, and 6-hydroxy-1-hexanethiol SAMs. Solvent vapor annealing followed by PEO removal led to the formation of nanopores with average diameters of 12 and 19 nm from the smaller and larger PS-hν-PEO, respectively. Cyclic voltammograms of 1,1'-ferrocenedimethanol showed that nanoporous films on cysteamine SAMs afforded nanopores reaching the underlying substrates at higher density than those on the other substrates. This result was attributed to balanced affinity of the cysteamine SAM surface with PS and PEO, which enhanced the vertical orientation of PEO microdomains. The generation of carboxyl groups associated with the photocleavage reaction was revealed by pH-dependent changes in the voltammogram of Fe(CN)63- that reflected electrostatic effects regulated by the protonation state of the carboxyl groups. The SAMs underneath the nanoporous films could be replaced by treatment with a thiol solution, as verified by voltammograms of l-ascorbic acid. These results suggest that thiolate SAM modification provides a simple means to control the interfacial orientation of PEO microdomains in thin PS-hν-PEO films.

Construction of Metal Organic Frameworks with Porphyrin on (110) Single Crystal Surfaces

Introduction These days, the use of photovoltaics such as solar cells have been expanding since they are clean, cost-effective, and carbon-free. The most common solar cells are crystalline silicon solar cells, but the efficiency is relatively low as about 20 %. The silicon absorbs only infrared radiation even though about half of the solar radiation is constituted by visible-light. On the other hand, porphyrin based solar cells show strong absorption in the visible region, so they have been increasing attention recently. Among them, porphyrin based dye sensitized solar cells (DSSCs) composed of semiconductors including TiO2 are investigated enthusiastically. Since DSSCs are the combination of dye which absorbs visible-light and TiO2 which absorbs ultraviolet radiation, they are thought to be effective. Porphyrin has a planar structure which is composed of four pyrrole subunits and has orientation dependent absorption. It absorbs visible-light most effectively irradiated from normal to the ring plane. Thus, it is important to fix the porphyrin ring flatly to the solid substrate surface to maximize the energy conversion efficiency. In order to control the orientation of molecules, metal organic frameworks (MOFs) which are highly crystalline porous coordination polymers can be utilized. MOFs are made from organic ligand and metal ion, and used as frameworks to introduce functional moiety such as porphyrin. MOFs can be fabricated by a layer-by-layer (LbL) fashion on solid substrate keeping the orientation and controlling the thickness and morphology without loss of bulk properties. As a linker between MOFs and substrates, self-assembled monolayers (SAMs) were employed to fix the MOFs on solid surface. The aim of this study is to construct the three-dimensional MOFs with porphyrin on single crystal Au(110) and TiO2(110) surfaces. As linker SAMs, cysteamine (CA) and isonicotinic acid (IA) SAMs were constructed on Au(110) and TiO2(110), respectively. On both the CA SAM / Au(110) and IA SAM / TiO2(110), Cu - TCPP (tetra-carboxyphenyl porphyrin) - 4,4’ bipyridine (bpy) MOF was fabricated by the LbL fashion. Each preparation process was structurally investigated by scanning tunneling microscopy (STM) and x-ray photoelectron spectroscopy (XPS). Experimental After annealing and quenching, Au(110) was immersed in 1 mM CA ethanol solution at room temperature for 1 hour to construct CA SAM on Au(110). The substrate was then successively soaked in 0.2 mM Cu(OAc)2 ethanol solution, 0.1 mM TCPP ethanol solution, and 0.2 mM bpy ethanol solution for 10 minutes, keeping the temperature of 40°C (Cu-TCPP-bpy MOF / CA SAM / Au(110)). After every immersion, the substrate was sonicated with ethanol for 10 min. In the same way, annealed TiO2(110) was soaked in saturated IA ethanol solution at 40°C for 1 day to construct IA SAM on TiO2(110). Subsequently, the substrate was immersed in 0.1 mM Cu(OAc)2 ethanol solution for 3, 5, and 10 minutes for comparison, keeping the temperature 40° (Cu2+ / IA SAM / TiO2(110)). The substrate was sonicated with ethanol for 10 min after every immersion. At each step, the structure and elemental composition were estimated by STM and XPS. Results and discussion In the case of Cu-TCPP-bpy MOF / CA SAM / Au(110), STM and XPS measurements were conducted. STM image reveals that CA SAM was homogeneously fabricated on the Au(110) surface. It was also observed by STM that Cu2+ and TCPP layers were constructed on the CA SAM with the LbL fashion, and bpy orientates vertical to the MOF sheet with porphyrin. Additionally, XP spectra in S2p and N1s indicates that the CA SAM was existed on Au(110), and those in Cu2p and N1s show that Cu2+, TCPP, and bpy were coordinated after each immersion. On the other hand, for Cu2+ / IA SAM / TiO2(110), XPS was performed. From a peak in N1s region, the modification of IA SAM on TiO2(110) was confirmed. Furthermore, the XP spectrum in Cu2p region shows the deposition of Cu2+ on IA SAM after the immersion.

Electrochemical Detection of Glucose by Using Glucose Oxidase Modified Platinum Electrode

In this research, the cysteamine self-assembled monolayers (SAM), is required to modify surface of pt electrode. Now, by using the method of chemical binding and electrochemical deposition the surface of platinum electrode was modified by glucose oxidase enzyme. Platinum plate (0.5cm×0.5cm), platinum rod and silver or silver chloride electrodes act as working, auxiliary and reference electrodes, individually. The manufactured of pt. /GOD electrode described by electrochemical impedance spectroscopy (EIS). The glucose oxidase modified platinum electrode was used for detection of glucose by Amperometric measurement. The amperometric response of the sensor was observed under some factors i.e. pH, Concentration, Temperature and applied potential. Detection of glucose can be taken by oxidation of enzymatically formed H2O2 at 0.8 V. A high current response was occur at pH 7.5 and potential +0.8 V. Enzyme is stronger at high temperature because of micro environment but at 28 °C biosensor show good response and this is working temperature. The stability of the enzyme electrode was also deliberated. Moreover, the performance of the glucose oxidase enzyme was reserved about 59.9% after a time of 15 days. These factors show pt/GOD electrode is good or successful biosensor for detection of glucose. Index Terms — Biosensor, Cysteamine self-assembled, Glucose oxidase, Glucose, Immobilization, DOI : 10.7176/CMR/11-4-03 Publication date : April 30 th 2019

Mechanistic Insights into Voltage-Driven Biocatalysis of a Cytochrome P450 Bactosomal Film on a Self-Assembled Monolayer

Simple construction of biocatalytically active films of cytochrome P450 (CYP) bactosomes is quite useful for low-cost, stereoselective, and nicotinamide adenine dinucleotide phosphate hydride-free drug metabolism assays, biosensing, and biocatalytic applications. We report here real-time monitoring of the formation of biocatalytically active films of membrane-bound human CYP 2C9 or 3A4 expressed with CYP reductase (CPR) in Escherichia coli (so-called bactosomes) on a cysteamine self-assembled monolayer of gold-infused quartz crystals. The CYP 2C9+CPR-containing bactosomes exhibited oxygen reduction currents and metabolite yields greater than those of the CYP 3A4+CPR film. The electrocatalytic property correlated with the greater levels of CPR activity and the amount of CYP 2C9 in the CYP 2C9+CPR bactosomes than in the CYP 3A4+CPR bactosomes. The electron mediating role of CPR in the CYP 2C9 bactosomal film (E°′ = −450 mV vs Ag/AgCl) toward electrocatalytic oxygen reduction and hydroxylation of diclofenac ...

Bactosomal Bioelectrode Design on a Cysteamine Self-Assembled Monolayer

We discuss here real-time monitoring of the formation of biocatalytically active films of membrane-bound human cytochrome P450 3A4 expressed with its reductase in E. Coli (so called bactosomes) on a cysteamine self-assembled monolayer of gold infused quartz crystals. Amount of immobilized P450 3A4 bactosomes was 471±21 μg cm–2 gold geometric area (N = 3 replicates). Scanning electron microscopic and impedance characterizations of the constructed P450 bactosomal films, electrocatalytic oxygen reduction activity, and voltage-driven (–0.6 V vs. Ag/AgCl, 1 M KCl) C-H functionalization of diclofenac to 4-hydroxydiclofenac typical of P450 3A4 catalysis are presented. The formation of 4-hydroxydiclofenac in the presence of added catalase (hydrogen peroxide scavenger) to the electrolysis solution suggested the reductase pathway of P450 3A4 catalysis followed in the designed bactosomal film, which is similar to the in vivo catalytic mechanism. This one-step construction of biocatalytically active films of cost-effective P450 bactosomes is quite useful for low cost, stereoselective, and NADPH-free drug metabolism assays, biosensing, and biocatalysis applications.

Boronate-Modified Interdigitated Electrode Array for Selective Impedance-Based Sensing of Glycated Hemoglobin

An impedance-based label-free affinity sensor was developed for the recognition of glycated hemoglobin (HbA1c). Interdigitated gold microelectrode arrays (IDAs) were first modified with a self-assembled monolayer of cysteamine followed by cross-linking with glutaraldehyde and subsequent binding of 3-aminophenylboronic acid (APBA), which selectively binds HbA1c via cis-diol interactions. Impedance sensing was demonstrated to be highly responsive to the clinically relevant HbA1c levels (0.1%-8.36%) with a detection and quantitation limit of 0.024% (3σ/slope) and 0.08% (10σ/slope), respectively. The specificity of the assay was evaluated with nonglycated hemoglobin (HbAo), showing that the impedance response remained unchanged over the concentration range of 10 to 20 g dL-1 HbAo. This demonstrated that the sensor system could be used to specifically distinguish HbA1c from HbAo. Moreover, the binding of HbA1c to the APBA-modified electrodes was reversible, providing a reusable sensing interface as well as showing a stable response after 4 weeks (96% of the initial response). When assaying normal (4.10%) and diabetic (8.36%) HbA1c levels (10 assays per day during a three-day period including a regeneration step after each assay), the overall assay reproducibility, expressed as relative standard error of the mean (n = 30), was 1.1%. The performance of the sensor system was also compared with a commercial method (n = 15) using patient-derived blood samples. A good agreement (Bland-Altman bias plot) and correlation (Passing-Bablok regression analysis) was demonstrated between the boronate-based affinity sensor and the standard method.

Ultrasensitive Nanoimmunosensor by coupling non-covalent functionalized graphene oxide platform and numerous ferritin labels on carbon nanotubes

An ultrasensitive electrochemical nanostructured immunosensor for a breast cancer biomarker carbohydrate antigen 15-3 (CA 15-3) was fabricated using non-covalent functionalized graphene oxides (GO/Py-COOH) as sensor probe and multiwalled carbon nanotube (MWCNTs)-supported numerous ferritin as labels. The immunosensor was constructed by immobilizing a monoclonal anti-CA 15-3 antibody on the GO modified cysteamine (Cys) self-assembled monolayer (SAM) on an Au electrode (Au/Cys) through the amide bond formation between the carboxylic acid groups of GO/Py-COOH and amine groups of anti-CA 15-3. Secondary antibody conjugated MWCNT-supported ferritin labels (Ab2-MWCNT-Ferritin) were prepared through the amide bond formation between amine groups of Ab2 and ferritin and carboxylic acid groups of MWCNTs. The detection of CA 15-3 was based on the enhanced bioelectrocatalytic reduction of hydrogen peroxide mediated by hydroquinone (HQ) at the GO/Py-COOH-based sensor probe. The GO/Py-COOH-based sensor probe and Ab2-MWCNT-Ferritin labels were characterized using cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), scanning electron microscope (SEM), transmission electron microscope (TEM), and x-ray photoelectron spectroscopy (XPS) techniques. Using differential pulse voltammetry (DPV) technique, CA 15-3 can be selectively detected as low as 0.01±0.07U/mL in human serum samples. Additionally, the proposed CA 15-3 immunosensor showed excellent selectivity and better stability in human serum samples, which demonstrated that the proposed immunosensor has potentials in proteomic researches and diagnostics.

Adjunction of Avidin to a Cysteamine Self-Assembled Monolayer for Impedimetric Immunosensor

In this work an impedimetric immunosensor based on affinity immobilization method of a biotin labelled anti-human IgG antibody, used as a model system, was reported. The experimental procedure involves the growth of a self-assembled monolayer of a thiol (cysteamine) carrying terminal amine groups on gold electrodes. Glutarardehyde, a homobifunctional cross-linker, was used as a coupling reagent for the covalent linking of avidin to the amine groups of cysteamine. The attachment of the biotin labeled antibodies (anti-Human IgG) to the subsequent modified gold electrode was achieved by affinity interactions tacking advantage of the strong avidin-biotin bridge. The stepwise assembly process of the electrode was interrogated by means of cyclic voltammetry, electrochemical impedance spectroscopy and contact angle measurements. The response of the antibody modified electrode to their target antigens was investigated in the presence of BSA (bovin serum albumin) in order to alleviate non-specific adsorption problems. A proposed electrical model was used to analyse the experimental data. The resulting immunosensor has a linear dynamic range of 100 - 900 ng?ml–1 of antigen and a detection limit of 100 ng?ml–1.

Supramolecular Control in Nanostructured Film Architectures for Detecting Breast Cancer.

The need for early detection of various diseases, including breast cancer, has motivated research into nanomaterials that can be assembled in organized films which serve as biosensors. Owing to the variety of possible materials and film architectures, procedures are required to design optimized biosensors. In this study, we combine surface-specific methods to monitor the assembly of antibodies on nanostructured films with two distinct architectures. In the first, a layer of the antibody type mouse anti-HER2 (clone tab250) was immobilized on a self-assembled monolayer (SAM) of 11-mercaptoundecanoic acid modified with N-hydroxysuccinimide (NHS) and 1-ethyl-3-(3-(dimethylamino)propyl)carbodiimide (EDC). In the second approach, a SAM of cysteamine was coated with a biotin/spreptavidin bilayer on which a layer of biotinylated antibody type MSx2HUp185/her biotin was adsorbed. The rougher, less passivating coating with cysteamine determined from cyclic voltammetry and scanning electron microscopy led to biosensors that are more sensitive to detect the breast cancer ERBB2 (HER2) biomarker in impedance spectroscopy measurements. This higher distinguishing ability of the cysteamine-containing film architecture was proven with information visualization methods to treat the impedance data. Polarization-modulated infrared reflection absorption spectroscopy (PM-IRRAS) confirmed that biosensing resulted from the antibody-ERBB2 antigen affinity.

Polyoxometalate nanostructured gold surfaces for sensitive biosensing of benzo[a]pyrene

We report the design of a polyoxometalate-nanostructured immunosensor for benzo[a]pyrene (B[a]P) detection. The organic–inorganic hybrid polyoxometalate (POM) (NBu4)3[PW11O39{(SiC6H4NH2)2O}] carrying two amine functions was covalently attached to a functionalized gold substrate to achieve a nanometric organization of amine groups at its surface. Pyrenebutyric acid (PBA) was subsequently grafted to amine groups to create the sensing layer. The detection of B[a]P in the indirect competitive format was carried out using a monoclonal anti-B[a]P antibody whose binding to the immunoprobe was monitored with a quartz crystal microbalance with dissipation measurement (QCM-D). The performances of the POM-nanostructured biosensor were compared to a reference sensor constructed from a cysteamine self-assembled monolayer. QCM-D measurements displayed significant input from POM-nanostructuration. Both the accessibility of the analogue on the surface and the analytical performances were enhanced showing a promising effect of this strategy of nanostructuration for the biosensing of small molecules.