Sensitive Detection Of C-reactive Protein Research Articles

Fabrication and sensing properties of a molecularly imprinted polymer on a photonic PDMS substrate for the optical detection of C-reactive protein

This work presents the development of a novel photonic-based sensor on a nanoscale patterned polydimethylsiloxane (PDMS) substrate acting as transducer, combined with a molecularly imprinted polymer for the rapid and sensitive detection of C-reactive protein (CRP). A silicon wafer was patterned using electron-beam lithography with a one-dimensional (1D) reflective grating (periodicity of about 400 nm) and then replicated using nanoimprint lithography. The latter silicon mould was used for replica moulding to generate a patterned PDMS. The unique 1D design gave PDMS photonic properties that were suitable as a substrate to assemble the biorecognition layer. The surface of the photonic PDMS was modified to become hydrophilic and additionally functionalized with vinyl silane groups to bind the imprinted polymer, resulting in a photonic molecularly imprinted polymer substrate (PMIPS). The developed method was successfully used to detect CRP in serum samples. The reflectance intensity decreased with increasing CRP concentrations, and the sensing PMIPS showed a linear response between 0.005 and 1.215 µg mL−1, a limit of detection of 0.003 µg mL−1, and a selective response to CRP when tested against serum calprotectin. Overall, this novel optical sensor can be used to detect CRP, a serum biomarker of inflammation, particularly in inflammatory bowel diseases.

One-Component Dual-Readout Aggregation-Induced Emission Nanobeads for Qualitative and Quantitative Detection of C-Reactive Protein at the Point of Care.

Fluorescent lateral flow immunoassay (LFA) systems are versatile tools for sensitive and quantitative detection of disease markers at the point of care. However, traditional fluorescent nanoparticle-based lateral flow immunoassays are not visible under room light, necessitate an additional fluorescent reader, and lack flexibility for different application scenarios. Herein, we report a dual-readout LFA system for the rapid and sensitive detection of C-reactive protein (CRP) in clinical samples. The system relied on the aggregation-induced emission nanobeads (AIENBs) encapsulated with red AIE luminogen, which possesses both highly fluorescent and colorimetric properties. The AIENB-based LFA in the naked-eye mode was able to qualitatively detect CRP levels as low as 8.0 mg/L, while in the fluorescent mode, it was able to quantitatively measure high-sensitivity CRP (hs-CRP) with a limit of detection of 0.16 mg/L. The AIENB-based LFA system also showed a good correlation with the clinically used immunoturbidimetric method for CRP and hs-CRP detection in human plasma. This dual-modal AIENB-based LFA system offers the convenience of colorimetric testing and highly sensitive and quantitative detection of disease biomarkers and medical diagnostics in various scenarios.

Electrochemiluminescence Aptasensor with Dual Signal Amplification by Silica Nanochannel-Based Confinement Effect on Nanocatalyst and Efficient Emitter Enrichment for Highly Sensitive Detection of C-Reactive Protein.

The rapid and sensitive detection of the important biomarker C-reactive protein (CRP) is of great significance for monitoring inflammation and tissue damage. In this work, an electrochemiluminescence (ECL) aptasensor was fabricated based on dual signal amplification for the sensitive detection of CRP in serum samples. The sensor was constructed by modifying a silica nanochannel array film (SNF) on a cost-effective indium tin oxide (ITO) electrode using the Stöber solution growth method. Gold nanoparticles (AuNPs) were grown in situ within the nanochannels using a simple electrodeposition method as a nanocatalyst to enhance the active electrode area as well as the ECL signal. The negatively charged nanochannels also significantly enriched the positively charged ECL emitters, further amplifying the signal. The recognition aptamer was covalently immobilized on the outer surface of SNF after modification with epoxy groups, constructing the aptasensor. In the presence of CRP, the formation of complexes on the recognitive interface led to a decrease in the diffusion of ECL emitters and co-reactants to the supporting electrode, resulting in a reduction in the ECL signal. Based on this mechanism, ECL detection of CRP was achieved with a linear range of 10 pg/mL to 1 μg/mL and a low limit of detection (7.4 pg/mL). The ECL aptasensor developed in this study offers advantages such as simple fabrication and high sensitivity, making promising applications in biomarker detection.

One-step paper-based SlipChip for the sensitive detection of C-reactive protein with porous platinum nanozyme-assisted signal amplification

The development of efficient and sensitive point-of-care testing is crucial for preparedness in the post-pandemic era. Although paper-based lateral flow assays have attracted attention and have various advantages for rapid, on-site diagnosis, they have low sensitivity. To overcome the limitations of the existing assays, in this study, we aimed to develop a new, one-step, nanozyme-amplified SlipChip for the sensitive detection of C-reactive protein (CRP). The SlipChip was constructed by combining wax-printed paper with different channel designs. The three-dimensional (3D) fluidic configuration of the SlipChip allowed for the sequential delivery of reagents, enabling mixing and signal amplification with a one-step sliding operation. As a signal-amplifying reagent, peroxidase-mimicking porous platinum nanozyme (pPtNZ) was synthesized using a simple wet chemical method. The pPtNZ conjugated on the test line catalyzes the oxidation of diaminobenzidine (DAB) in the presence of hydrogen peroxide, increasing the color intensity. The immunoassay results of the SlipChip were easily interpreted within 20 min, and the color intensity was visually enhanced by DAB precipitation over time, resulting in up to 6-fold signal amplification. The proposed pPtNZ-SlipChip exhibited high analytical performances for the one-step detection of serum and salivary CRP from 0.1 to 1000 ng/mL, with a limit of detection of 0.03 ng/mL. These results revealed the potential and applicability of the pPtNZ-SlipChip, with the advantages of simplicity, sensitivity, low cost, and portability for on-site detection and point-of-care testing.

Photoelectrochemical aptasensor based on PTCDA/TiO2 efficient sensitization effect for sensitive protein analysis

Herein, a photoelectrochemical (PEC) aptasensor for sensitive detection of C-reactive protein (CRP) was designed based on the efficient sensitization effect of 3,4,9,10-Perylene tetracarboxylic dianhydride (PTCDA) towards titanium dioxide (TiO2). Initially, TiO2 and PTCDA were continuously incubated on electrode surface to produce a strong PEC signal owing to the PTCDA/TiO2 sensitization structure. Subsequently, electrodeposition of gold nanoparticles (dep Au) was performed to connect with the incubated CRP aptamer (S1) via AuN bonds. After that, 1-hexanethiol (HT) was added on the surface of the modified electrode to block the non-specific adsorption sites. Eventually, with the addition of CRP, the PEC signal decreased significantly because the CRP-S1 complexes generated by the specific recognition of CRP and S1 resulted in high steric hindrance effect and inhibited electron transfer. Under optimal conditions, the proposed PEC aptasensor was applied successfully for sensitive CRP detection with a wide linear range from 10 pg/mL to 10 μg/mL and a low detection limit of 3.33 pg/mL. Moreover, the designed PEC aptasensor based on PTCDA/TiO2 sensitized structure might be a versatile tool that provides a powerful alternative to traditional proteins, ions and RNAs detection methods by changing the corresponding recognition sequences.

Water soluble silicon quantum dot based fluorescence immunoassay probe for C-reactive protein (CRP) detection

Silicon quantum dot (SiQD) based water soluble fluorescence immunoassay probe with aggregation induced emission features (AIE) was developed for the sensitive detection of C-reactive protein (CRP) biomarker. The solid state and solution phase photophysical properties of SiQD attributed from AIE effect was studied successfully. Later the ability of CRP antibody functionalized SiQD against the detection CRP antigen was evaluated systematically. The method relies on the decrease in fluorescence intensity of CRP antibody functionalized SiQD (CRPAb) upon the gradual addition of CRP antigen (CRPAg) via antigen-antibody interaction. The intensity of the antibody functionalized fluorophore quenched linearly by the addition of CRPAg within the concentration range of 50–150 nM and the limit of detection (LOD) was as sensitive as about 0.89 nM. Detection mechanism was assigned further by evaluating the interaction that exists between SiQD-CRPAb and CRPAg, which suggests that the effective complexation through a lock and key mechanism could be the plausible reason behind the fluorescence quenching of SiQD-CRPAb in presence of CRPAg. The solid state fluorescence features of SiQD arises from AIE phenomena which suggest that as a promising alternative to conventional QDs and organic fluorophores, integration of SiQD with lateral flow strips could result in powerful eco-friendly point-of-care device for in vitro diagnosis and the detection of other biomarkers.

Faradaic electrochemical impedimetric analysis on MoS2/Au-NPs decorated surface for C-reactive protein detection

BackgroundA label-free Faradaic electrochemical impedimetric was developed for a highly sensitive detection of C-reactive protein using a gold interdigitated microelectrode bio-sensing platform enhanced by a gold nanoparticle-decorated molybdenum disulfide (Au-NPs/MoS2) nanosheet via selected chemical linking processes. C-reactive protein (C-RP), a crystalline protein, generates by the liver and hikes when there is inflammation throughout the patients’ body. The concentrations of C-RP plasma levels tend to increase rapidly when the patient facing major injury which will lead to cardiovascular disease (CVD). MethodsThe 5 µm microelectrode and gap size g-IDE with the nanostructured materials was demonstrated to increase the impedimetric detection response in Faradaic-mode electrochemical impedance spectroscopy high performance detection environment. The high surface area-to-volume ratio of the modified Au-NPs/MoS2 nanocomposite increased the probes loading onto the transducer and enhanced the impedimetric detection response of the C-RP target post-binding due to an amplified net change in the charge transfer resistance. The developed immunoassay revealed a linear detection of C-RP biomarker in a logarithmic-scale from as low as 1 fg/mL up to 1 µg/mL, and a limit of detection of 0.01 fg/mL. The sensor shows great potential as an early warning risk for cardiovascular disease at a threshold concentration value of C-RP at 1 µg/mL. Significant findingsThe biosensor demonstrates an excellent discrimination against other competing proteins in serum, exhibiting the highest predilection to C-RP spiked human serum target. The sensor's reproducibility is reported within an acceptable range of relative standard deviation of 1–4% for n = 3.

Wave-shaped microfluidic chip assisted point-of-care testing for accurate and rapid diagnosis of infections

BackgroundEarly diagnosis and classification of infections increase the cure rate while decreasing complications, which is significant for severe infections, especially for war surgery. However, traditional methods rely on laborious operations and bulky devices. On the other hand, point-of-care (POC) methods suffer from limited robustness and accuracy. Therefore, it is of urgent demand to develop POC devices for rapid and accurate diagnosis of infections to fulfill on-site militarized requirements.MethodsWe developed a wave-shaped microfluidic chip (WMC) assisted multiplexed detection platform (WMC-MDP). WMC-MDP reduces detection time and improves repeatability through premixing of the samples and reaction of the reagents. We further combined the detection platform with the streptavidin–biotin (SA-B) amplified system to enhance the sensitivity while using chemiluminescence (CL) intensity as signal readout. We realized simultaneous detection of C-reactive protein (CRP), procalcitonin (PCT), and interleukin-6 (IL-6) on the detection platform and evaluated the sensitivity, linear range, selectivity, and repeatability. Finally, we finished detecting 15 samples from volunteers and compared the results with commercial ELISA kits.ResultsDetection of CRP, PCT, and IL-6 exhibited good linear relationships between CL intensities and concentrations in the range of 1.25–40 μg/ml, 0.4–12.8 ng/ml, and 50–1600 pg/ml, respectively. The limit of detection of CRP, PCT, and IL-6 were 0.54 μg/ml, 0.11 ng/ml, and 16.25 pg/ml, respectively. WMC-MDP is capable of good adequate selectivity and repeatability. The whole detection procedure takes only 22 min that meets the requirements of a POC device. Results of 15 samples from volunteers were consistent with the results detected by commercial ELISA kits.ConclusionsWMC-MDP allows simultaneous, rapid, and sensitive detection of CRP, PCT, and IL-6 with satisfactory selectivity and repeatability, requiring minimal manipulation. However, WMC-MDP takes advantage of being a microfluidic device showing the coefficients of variation less than 10% enabling WMC-MDP to be a type of point-of-care testing (POCT). Therefore, WMC-MDP provides a promising alternative to POCT of multiple biomarkers. We believe the practical application of WMC-MDP in militarized fields will revolutionize infection diagnosis for soldiers.

A zirconium-based metal-organic framework sensitized by thioflavin-T for sensitive photoelectrochemical detection of C-reactive protein.

Herein, a novel photoelectrochemical (PEC) assay was developed for the sensitive detection of C-reactive protein (CRP) based on a zirconium-based metal-organic framework (PCN-777) as the photoelectric material and thioflavin-T (Th-T) as the effective signal sensitizer coupled with rolling circle amplification (RCA).

Detection of C-reactive protein on an integrated microfluidic system by utilizing field-effect transistors and aptamers.

Cardiovascular diseases (CVDs) cause more than 17 × 106 deaths worldwide on a yearly basis. Early diagnosis of CVDs is therefore of great need. The C-reactive protein (CRP) is an important biomarker for analyzing the risks of CVDs. In this work, CRP-specific aptamers with high sensitivity and specificity and field-effect-transistor (FET) devices were used to recognize and detect CRP by using an integrated microfluidic system automatically while consuming less volumes of reagents and samples (about 5 μm). In order to package the FET device into the microfluidic chip, a new method to prevent liquid leakage was proposed. Sensitive detection of CRP has been demonstrated on the developed microfluidic system. It is the first time that aptamer-FET assays could be realized on an integrated microfluidic system. Experimental results showed that the aptamer-FET assay was capable of detecting CRP with concentrations ranging from 0.625 mg/l to 10.000 mg/l, which may be promising for early diagnosis of CVDs.

The improved sensitive detection of C-reactive protein based on the chemiluminescence immunoassay by employing monodispersed PAA-Au/Fe3O4 nanoparticles and zwitterionic glycerophosphoryl choline.

Nanocomposites are emerging complex nanoscale materials composed of different phases that have received considerable attention as a result of their widespread impact on a myriad of fields. Having said this, it is important to recognize that poor colloidal stabilities in physiological media and non-specific interactions with biology are key limitations prohibiting their successful translation into the development of practical technologies with real impact. Herein, PAA coated Au/Fe3O4 composite magnetic nanoparticles (NPs) with monodispersity and high biostability were engineered for the development of a highly sensitive C-reactive protein (CRP) chemiluminescence immunoassay (CLIA), based on the horseradish peroxidase (HRP)-luminol-H2O2 system. In addition, zwitterionic glycerophosphoryl choline (GPC) was employed as a co-blocking agent to reduce the nonspecific adsorption. This assay reports a limit of detection (LOD) of 2.5 pM with high reproducibility verified clinically and demonstrating improved progress towards translation into clinics.

Highly sensitive detection of C-reactive protein using a novel dissolution approach in a dye-doped silica nanoparticle-based fluorescence immunoassay

A novel dissolution approach was used to achieve enhanced sensitivity in a nanoparticle (NP) immunoassay for the detection of a cardiovascular disease marker, C-reactive protein (CRP).

A plasmonic chip-based bio/chemical hybrid sensing system for the highly sensitive detection of C-reactive protein.

A synthetic polymer ligand-grafted plasmonic chip was fabricated and demonstrated a highly sensitive detection of C-reactive protein (CRP) by grating-coupled surface plasmon field-enhanced fluorescence. Poly(2-methacryloyloxyethyl-phosphorylcholine) was used as a CRP-specific polymer ligand layer and was grafted onto the plasmonic chip using surface-initiated controlled/living radical polymerization (limit of detection: ca. 10 pM).

Direct Covalent Biomolecule Immobilization on Plasma-Nanotextured Chemically Stable Substrates.

A new method for direct covalent immobilization of protein molecules (including antibodies) on organic polymers with plasma-induced random micronanoscale topography and stable-in-time chemical functionality is presented. This is achieved using a short (1-5 min) plasma etching and simultaneous micronanotexturing process, followed by a fast thermal annealing step, which induces accelerated hydrophobic recovery while preserving important chemical functionality created by the plasma. Surface-bound biomolecules resist harsh washing with sodium dodecyl sulfate and other detergents even at elevated temperatures, losing less than 40% of the biomolecules bound even at the harshest washing conditions. X-ray photoelectron spectroscopy, secondary-ion mass spectrometry, and electron paramagnetic resonance are used to unveil the chemical modification of the plasma-treated and stabilized surfaces. The nanotextured and chemically stabilized surfaces are used as substrates for the development of immunochemical assays for the sensitive detection of C-reactive protein and salmonella lipopolysaccharides through immobilization of the respective analyte-specific antibodies onto them. Such substrates are stable for a period of 1 year with ambient storage.

Sensitive detection of C-reactive protein in serum by immunoprecipitation–microchip capillary gel electrophoresis

Sepsis represents a significant cause of mortality in intensive care units. Early diagnosis of sepsis is essential to increase the survival rate of patients. Among others, C-reactive protein (CRP) is commonly used as a sepsis marker. In this work we introduce immune precipitation combined with microchip capillary gel electrophoresis (IP–MCGE) for the detection and quantification of CRP in serum samples. First high-abundance proteins (HSA, IgG) are removed from serum samples using affinity spin cartridges, and then the remaining proteins are labeled with a fluorescence dye and incubated with an anti-CRP antibody, and the antigen/antibody complex is precipitated with protein G-coated magnetic beads. After precipitation the complex is eluted from the beads and loaded onto the MCGE system. CRP could be reliably detected and quantified, with a detection limit of 25ng/μl in serum samples and 126pg/μl in matrix-free samples. The overall sensitivity (LOQ=75ng/μl, R2=0.9668) of the method is lower than that of some specially developed methods (e.g., immune radiometric assay) but is comparable to those of clinically accepted ELISA methods. The straightforward sample preparation (not prone to mistakes), reduced sample and reagent volumes (including the antibodies), and high throughput (10 samples/3h) are advantages and therefore IP–MCGE bears potential for point-of-care diagnosis.

Sensitive detection of C-reactive protein using optical fiber Bragg gratings

An accurate and highly sensitive sensor platform has been demonstrated for the detection of C-reactive protein (CRP) using optical fiber Bragg gratings (FBGs). The CRP detection has been carried out by monitoring the shift in Bragg wavelength (ΔλB) of an etched FBG (eFBG) coated with an anti-CRP antibody (aCRP)-graphene oxide (GO) complex. The complex is characterized by Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy and atomic force microscopy. A limit of detection of 0.01mg/L has been achieved with a linear range of detection from 0.01mg/L to 100mg/L which includes clinical range of CRP. The eFBG sensor coated with only aCRP (without GO) show much less sensitivity than that of aCRP–GO complex coated eFBG. The eFBG sensors show high specificity to CRP even in the presence of other interfering factors such as urea, creatinine and glucose. The affinity constant of ∼1.1×1010M−1 has been extracted from the data of normalized shift (ΔλB/λB) as a function of CRP concentration.

One-step kinetics-based immunoassay for the highly sensitive detection of C-reactive protein in less than 30 min

This article reveals a rapid sandwich enzyme-linked immunosorbent assay (ELISA) for the highly sensitive detection of human C-reactive protein (CRP) in less than 30min. It employs a one-step kinetics-based highly simplified and cost-effective sandwich ELISA procedure with minimal process steps. The procedure involves the formation of a sandwich immune complex on capture anti-human CRP antibody-bound Dynabeads in 15min, followed by two magnet-assisted washings and one enzymatic reaction. The developed sandwich ELISA detects CRP in the dynamic range of 0.3 to 81ngml−1 with a limit of detection of 0.4ngml−1 and an analytical sensitivity of 0.7ngml−1. It detects CRP spiked in diluted human whole blood and serum with high analytical precision, as confirmed by conventional sandwich ELISA. Moreover, the results of the developed ELISA for the determination of CRP in the ethylenediaminetetraacetic acid plasma samples of patients are in good agreement with those obtained by the conventional ELISA. The developed immunoassay has immense potential for the development of rapid and cost-effective in vitro diagnostic kits.

Label-free detection of C-reactive protein using a carbon nanofiber based biosensor

We report the sensitive detection of C-reactive protein (CRP), a biomarker for cardiac disease, using a carbon nanofiber based biosensor platform. Vertically aligned carbon nanofibers were grown using plasma enhanced chemical vapor deposition to fabricate nanoelectrode arrays in a 3×3 configuration. Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used for the CRP detection. The CV responses show a 25% reduction in redox current upon the immobilization of anti-CRP on the electrode where as a 30% increase in charge transfer resistance is seen from EIS. Further reduction in redox current and increase in charge transfer resistance result from binding of CRP on anti-CRP immobilized surface, proportional to the concentration of the CRP target. The detection limit of the sensor is found to be ~90pM or ~11ng/ml, which is in the clinically relevant range. Control tests using non-specific myoglobin antigen confirmed the specificity of the present approach.

Wash-free highly sensitive detection of C-reactive protein using gold derivatised triangular silver nanoplates

Rapid, wash-free highly sensitive detection of C-reactive protein is reported using nanoplate biosensor sols.

Sensitive detection of cardiac biomarker using ZnS nanoparticles as novel signal transducers

C-reactive protein (CRP), a 115 kDa pentameric protein, is one of the important cardiac biomarkers that are indicative of coronary heart events. Sensitive detection of CRP in human serum is critical for the diagnosis of coronary heart disease. This work presents a sensitive sandwich immunoassay for the detection of CRP in human serum using zinc sulfide (ZnS) nanoparticles as novel fluorescence signal transducers. In this assay, monoclonal anti-CRP antibodies are used to capture CRP in human serum, and then the captured CRPs are incubated with biotinylated monoclonal anti-CRP and Neutravidin coated ZnS nanoparticle to form sandwich immunocomplexes. Quantification of CRP occurs when zinc ions released from ZnS nanoparticle labels are mixed with zinc-ion sensitive fluorescence indicator Fluozin-3 for fluorescence generation. The developed assay presents a detection limit around 10 pM and a detection range with more than two orders of magnitude.