Antigen Immunoassay Research Articles

A voltammetric immunoassay for the carcinoembryonic antigen using silver(I)-terephthalate metal-organic frameworks containing gold nanoparticles as a signal probe

A voltammetric immunoassay forthe carcinoembryonic antigen (CEA) was developed using silver metal-organic frameworks (Ag-MOFs) as a signal probe. The Ag-MOFs contained a substantial amount of Ag(I) whose electrochemical signal was relatively stable. Therefore, the Ag-MOFs were viable signal probes. The signal can be detected without previous acid dissolution and preconcentration steps. This simplifies the detection steps and reduces the detection time. If CEA binds to its antibody on the modified electrode, the signal for Ag(I) becomes reduced. Measurements were best performed at a potential as low as 0.07V (vs. SCE) which made the electrode hardly sensitive to potential electroactive interferents. Under optimized conditions, the method included a wide linear response range (0.05 to 120ngmL-1) and a low detection limit of 8.0fgmL-1 (S/N = 3). The synthesis of Ag-MOFs is relatively simple, and Ag-MOFs are not only used as nanocarriers for immobilizing anti-CEA but also acted as electroactive materials for a signal probe. The voltammetric immunoassay is stable, inexpensive, sensitive, and selective. Based on these advantages, the method holds great promise for applications in the point-of-care disease and environmental monitoring. Graphical abstract Schematic presentation of a novel electrochemical immunoassay for thecarcinoembryonic antigen (CEA) using silver metal-organic frameworks (Ag-MOFs) as a signal probe.

False-positive Candida and Aspergillus antigen testing in recipients of allogeneic haematopoietic cell transplantation due to administration of parenteral nutrition and fixed combinations of piperacillin-tazobactam.

False positivity of antigen immunoassays used as an early diagnostic tool to detect invasive fungal infections is known. Interpretation of the assay needs the identification of sources which could affect the specificity of the test. We focused on the influence of parenteral nutrition (PN) and piperacillin-tazobactam (TZP) on fungal immunoassays. Measurable amounts of Candida antigen mannan were detected in several compounds of PN and TZP in a previous in vitro study. In the current study, 84 patients undergoing allogeneic haematopoietic cell transplantation receiving either TZP, PN or both were monitored with Aspergillus and Candida antigen assay. Six patients were analysed closer in a kinetic analysis with more frequent blood sampling to detect mannan. PN in diverse compositions as well as TZP did not increase significantly the amount of mannan and the Aspergillus antigen in serum. We could not confirm the positive results of the in vitro study. Physicians should be aware that mannan antigenemia due to drug infusion could be a transient issue and should be considered in the interpretation of fungal immunoassays, although we could not find clinically relevant effects on mannan levels.

PH responsive amperometric immunoassay for carcinoma antigen 125 based on hollow polydopamine encapsulating methylene blue

A sandwich-type amperometric immunosensor based on pH responsive sensitivity amplifiable route was elaborately designed for determination of carcinoma antigen 125 (CA 125). The sensitivity of amperometric immunosensor depends on a change of current response (ΔI) attributed to antigen to be tested per unit concentration. Employing zeolitic imidazolate framework (ZIF-8) as the template, the hollow-structured polydopamine framework with high specific surface area can be fabricated and it can enrich large amounts of signal molecules. The pH responsive hollow polydopamine composites which loaded methylene blue (MB) covalently coupled with antibodies (Ab2) were implemented as smart labels and the Au-rGO with high absorption capacity was served as substrate to capture signal molecules. Different from traditional electrochemical immunosensor, HCl led to the destruction of the labels and disassociation of antibody-antigen interaction layer. The impedance of modified interface was dramatically decreased and the distance between signal molecules and the sensing interface was shortened, giving rise to the improvement of electron transfer efficiency and amplification of current response. With the help of pH responsive strategy, ΔI caused by antigen was effectively elevated, leading to the sensitivity amplification of immunosensor. Under optimum conditions, the amperometric immunosensor with good reproducibility, high sensitivity and specificity exhibited an ultralow detection limit of 0.336 μU mL−1 (S/N = 3) and a wide linear range from 0.0001 to 100 U mL−1 for CA 125 detection.

Diagnostic and analytical performance of the hepatitis B core related antigen immunoassay in hepatitis B patients

BackgroundNovel serological markers for Hepatitis B virus (HBV) infection are needed for prognosis and guidance of therapy. ObjectiveWe evaluated the diagnostic performance of the Fujirebio Lumipulse G HBcrAg immunoassay on the Fujirebio LUMIPULSE G1200 analyzer. Study designAnalytical performance was examined using three HBeAg positive HBV samples. Diagnostic specificity was assessed using subpanels of 54 confirmed acute HAV, HCV, HEV, B19, CMV and EBV infections. Diagnostic sensitivity was investigated in well-defined HBV positive patient groups, both treated and untreated, including immunocompromised patients. ResultsThe Lumipulse G HBcrAg immunoassay provided a linear measurement at a dilution between 1:100 and1:10,000. Six out of 54 samples showed non-specific reactivity in sera from acute CMV, EBV and HEV infections, of which 2 of them >3 log U/ml. The highest levels of HBcrAg were measured in HBeAg positive patients, in both treated and untreated as well as in immunocompromised patients. Untreated patients had relatively low serum HBcrAg levels in the inactive carrier phase, which increased upon progression into the HBeAg-negative hepatitis phase. Also, we showed that the applicability of HBcrAg to distinguish between patients with resolved HBV infection and false-positive reactivity to solitary anti-HBc is limited. ConclusionsOur study demonstrated significant differences in HBcrAg levels depending on HBeAg status, the clinical phase, as well as the treatment status. Specificity of the assay is good; only 2 out of 54 samples showed reactivity above 3 log U/ml. Before implementing the assay in clinical practice, additional research in larger patient cohorts should be carried out.

A new visual immunoassay for prostate-specific antigen using near-infrared excited CuxS nanocrystals and imaging on a smartphone.

A photothermal immune-imaging assay was innovatively designed for the visual quantitative detection of cancer biomarkers by coupling CuxS nanocrystals with a portable infrared thermal imager on a smartphone. The rolling circle amplification (RCA) technique was used for the formation of a CuxS nanocrystal concatemer, thus opening up new territories in immunoassay development.

Determination of alkaline phosphatase activity and of carcinoembryonic antigen by using a multicolor liquid crystal biosensor based on the controlled growth of silver nanoparticles

An ultrasensitive liquid crystal biosensor is described for multicolor visualization of the activity of alkaline phosphatase (ALP) based on the controlled growth of silver nanoparticles. The enzymatic product is accumulated on the surface of the LC sensing film by means of silver deposition, and the birefringent signal (observed with a polarizing microscope) is strongly enhanced as a result. The presence of AuNPs also enhances the sensitivity by about 4 orders of magnitude. The bright spots in polarized optical microscopy (POM) images increase with increasing activities of ALP. The signal intensities of the spots are then calculated by using Photoshop software and by multiplying the average brightness of the spots by the pixel value. The detection limit for ALP is 1.2 nU·mL-1, which is 5-7 orders of magnitude lower than other colorimetric or fluorometric methods. The method was applied to a highly sensitive immunoassay for the carcinoembryonic antigen (CEA) by integrating immunomagnetic separation. The immunoassay was applied to the analysis of complex samples without tedious sample pretreatment, and a detection limit as low as 0.35pg·mL-1 of CEA was achieved. The method has attractive features in that it provides an ultrasensitive multicolor visualization approach for enzymes such as ALP, but also paves the way to a new kind of immunoassay coupled to immunomagnetic separation. Graphical abstract A signal enhanced liquid crystal (LC)-based multicolor immunosensor is describedthat is based on immunomagnetic separation and biometallization. Alkaline phosphatase (ALP) and carcinoembryonic antigen (CEA) can be easily visualized by bare eyes using the polarized optical microscopy (POM) images of LCs.

Switchable fluorescence immunoassay using gold nanoclusters anchored cobalt oxyhydroxide composite for sensitive detection of imidacloprid

The fabrication of a convenient fluorescence-based immunoassay (FIA) for specific antigen attract increasing concern but remains a considerable challenge, especially the laborious synthesis of nanomaterials-antibody conjugates. Herein, we circumvent this drawback by introducing a specific fluorescence signal response procedure to commercially available alkaline phosphatase (ALP)-labeled conventional immunoassay for imidacloprid detection. In this FIA protocol, gold nanoclusters (AuNCs) can anchor on the surface of two-dimensional cobalt oxyhydroxide (CoOOH) nanoflakes to form nanocomposite, resulting in remarkable decrease of fluorescence intensity. The quenching effects can be effectively reversed by introducing ascorbic acid that can trigger the decomposition of CoOOH nanoflakes. Notably, the corresponding fluorescence response induced by ascorbic acid was related to ALP activities labeled on antibody. After competitive immunoreaction, the ALP-labeled antibodies were bounded to immobilized antigen, which can regulate the fluorescence change. Utilizing fluorescence switching of system, the 50% inhibition concentration (IC50) value of FIA toward imidacloprid was obtained at 1.3 ng mL−1 which was 60-fold-of-magnitude more sensitive than that of conventional ELISA (86.4 ng mL−1). This FIA protocol not only develops new prospects for pesticide detection, but also opens up potent strategy of fluorogenic immunoassay.

High-sensitivity lateral flow immunoassay with a fluorescent lanthanide nanoparticle label

Lateral flow (LF) immunoassays are commonly used for point-of-care testing and typically incorporate visually read reporters, such as gold particles. To improve sensitivity and develop quantitative LF immunoassays, visual reporters can be replaced by fluorescent reporters detected by an instrument. In this study, we used fluorescent europium(III) chelate doped nanoparticle (Eu-np) reporters to develop a quantitative high-sensitivity LF immunoassay for free prostate specific antigen (fPSA). Furthermore, we tested different simplified formats of the assay and the effect of different modifiable parameters on the detection limit of the assay: dynamic range, assay duration and number of assay steps. The molar detection limits of the different assay formats were compared with published detection limits of LF immunoassays with different reporters. The cutoff was calculated from 11 female serum samples. The detection limit of the sensitivity optimized fPSA assay with fPSA spiked into pooled female serum was 0.01 ng/ml, which is approximately 100-fold lower than the most sensitive gold particle LF assays and 10-fold lower than other Eu-np and carbon nanoparticle based LF immunoassays. Thus, Eu-np reporters can be used to develop highly sensitive and quantitative LF immunoassays.

How to Improve Sensitivity of Sandwich Lateral Flow Immunoassay for Corpuscular Antigens on the Example of Potato Virus Y?

A simple approach was proposed to decrease the detection limit of sandwich lateral flow immunoassay (LFIA) by changing the conditions for binding between a polyvalent antigen and a conjugate of gold nanoparticles (GNPs) with antibodies. In this study, the potato virus Y (PVY) was used as the polyvalent antigen, which affects economically important plants in the Solanaceae family. The obtained polyclonal antibodies that are specific to PVY were characterized using a sandwich enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance (SPR). For LFIA, the antibodies were conjugated with GNPs with a diameter of 17.4 ± 1.0 nm. We conducted LFIAs using GNP conjugates in a dried state on the test strip and after pre-incubation with a sample. Pre-incubating the GNP conjugates and sample for 30 s was found to decrease the detection limit by 60-fold from 330 ng∙mL−1 to 5.4 ng∙mL−1 in comparison with conventional LFIA. The developed method was successfully tested for its ability to detect PVY in infected and uninfected potato leaves. The quantitative results of the proposed LFIA with pre-incubation were confirmed by ELISA, and resulted in a correlation coefficient of 0.891. The proposed approach is rapid, simple, and preserves the main advantages of LFIA as a non-laboratory diagnostic method.

Amplified photoelectrochemical immunoassay for the tumor marker carbohydrate antigen 724 based on dye sensitization of the semiconductor composite C3N4-MoS2.

The authors describe an amplified photoelectrochemical immunoassay for the tumor marker carbohydrate antigen 724 (CA724). The method employs a C3N4-MoS2 semiconductor as the photoelectric conversion layer. The nanocomposite was characterized by transmission electron microscopy, Fourier transform infrared spectroscopy, X-ray powder diffraction, and UV-vis diffuse reflectometry. The dye eosin Y was encapsulated into CaCO3 nanospheres which then were used as labels for antibody against CA724. In addition,Fe3O4 nanospheres were employed as magnetic platform for constructing photoelectrochemical sandwich immunoassay. TheCaCO3 nanospheres can be dissolved with aid of ethylene diamine tetraacetic acid (EDTA) and the carried eosin Y in CaCO3 is released. The released dyes sensitizes the C3N4-MoS2 semiconductor, which induces photocurrent amplification. Under optimal conditions and at a typical working voltage of 0V (vs. SCE), the photocurrent increases linearly in the range of 0.05mUmL-1 to 500mUmL-1 ofCA724, with a 0.02mUmL-1 detection limit. Graphical abstract TheC3N4-MoS2 complex, with high efficiency of electron transport, was synthesized to construct aphotoelectrochemical analyticalplatform. A sandwich-type immunoassay was established on the surface of magnetic beads. Carbohydrate antigen 724 in sample was detected sensitively by using sensitization of released eosin Y as signal amplifiery.

Aerogels prepared from polymeric β-cyclodextrin and graphene aerogels as a novel host-guest system for immobilization of antibodies: a voltammetric immunosensor for the tumor marker CA 15-3.

A three-dimensional porous network graphene aerogel (GAs) with large specific area and excellent conductivity was loaded with β-cyclodextrin polymer (Pβ-CD) to serve as a support for immobilization of antibodies. A highly sensitive immunosensor for the cancer marker carbohydrate antigen 15-3 (CA15-3) was designed based on the use of Pβ-CD/GAs. The large specific area of GAs warrants high loading with antibodies, and their excellent electrical conductivity warrants strong electrical signals. Based on the synergistic effect of GAs and Pβ-CD, an immunoassay was designed that is making use of hexacyanoferrate as an electrochemical probe and having a pleasantly low working potential of 0.2V (vs. SCE). Response is linear in the 0.1mUmL-1 to 100UmL-1 activity range, and the lower detection limit is 0.03mUmL-1 (at S/N= 3). The immunoassay is stable, selective and reproducible. It was applied to the analysis of spiked samples, and results were satisfactory. Graphical abstract Schematic of an electrochemical immunoassay for the carbohydrate antigen 15-3. It is based on the use of β-cyclodextrin polymer and a graphene aerogel.

Localized surface plasmon resonance biosensor using nanopatterned gold particles on the surface of an optical fiber

Fiber-optic localized surface plasmon resonance (FO LSPR) sensors are fabricated by immobilizing metal nanoparticles on the surface of an optical fiber, and have advantages in their miniaturization, simple optical set-up, low cost, and remote sensing. However, the conventional FO LSPR sensor based on chemically synthesized metal nanoparticles lacks durability and fabrication reproducibility, due to the rearrangement and irregular sizes of metal nanoparticles. In this paper, in order to overcome these problems, focused ion beam (FIB) milling is utilized to pattern nanostructures from a metallic film deposited on the end-face of an optical fiber. We also present a design for the fabrication processes to utilize multi-mode fibers. The proposed method is evidenced by measuring the output intensities of an FO LSPR sensor for various refractive index solutions. To verify fabrication reproducibility, sensors fabricated in different batches are compared. Finally, a prostate-specific antigen (PSA) immunoassay is performed to verify the possibility of the fabricated sensor system as a biosensor. PSA is the most useful biomarker for the screening, early detection, and follow up of prostate cancer. Any change in the low PSA levels is important to detect early diagnosis and recurrence of prostate cancer. We measure the PSA to 0.1 pg/ml using the proposed FO LSPR sensor.

Within-host evolution of virus variants during chronic infection with novel GII.P26-GII.26 norovirus

BackgroundNoroviruses are a leading cause of acute gastroenteritis in all age groups. They generally cause a rapidly self-limiting illness. However, chronic norovirus diarrheal disease occurs in immunocompromised individuals, and is accompanied by persistent shedding of infectious norovirus in stool. ObjectivesThe study aims to characterize a novel GII.P26-GII.26 norovirus strain. Furthermore, it analyses viral mutations arising during chronic infection of an immunocompromised host. Study designOver the course of more than three years, stool samples were obtained from an immunocompromised patient and screened for the presence of norovirus RNA by real-time PCR and norovirus antigen by immunoassay. Viral population kinetics was analyzed by conventional and high-throughput-sequencing. ResultsReal-time PCR yielded high amounts of norovirus RNA in the stool, but antigen immunoassays failed to detect the virus. The near complete norovirus genome was assigned as novel GII.P26-GII.26 genotype. Conventional as well as high-throughput sequencing pointed to a heterogeneous viral population with low rates of non-synonymous substitutions. Within-host evolution was enhanced in non-structural protein p22 and the N-terminal arm of the capsid protein VP1 but reduced in the viral polymerase RdRp. Intermittent non-synonymous substitutions in the protruding domain of the VP1 reverted fully over time. ConclusionsConfirmation of novel GII.P26-GII.26 norovirus genotypes provides insight into norovirus genetic diversity. The study further illustrates norovirus infection as an important differential diagnosis of recurrent persistent diarrhea in immunocompromised patients. The provided data on within-host evolution contribute to the insight of the mechanisms of viral persistence and pathogenesis in chronic norovirus infections.

Hepatitis C core antigen highly correlated to HCV RNA

Hepatitis C virus core antigen (HCV-Ag) immunoassay has been proposed as a more cost and time efficient one-step alternative to the current two-step screening and diagnostic process. This study investigates the correlation between the HCV-Ag immunoassay and the current gold standard of Hepatitis C Virus (HCV) ribonucleic acid (RNA) molecular assay. Stored sera of 221 consecutive treatment-naive patients tested anti-HCV positive were selected to undergo both HCV-Ag immunoassay and HCV RNA molecular assay. Active infection status and HCV genotype were determined using both assays, and correlation was calculated using a logarithmic scale. Among 221 anti-HCV-positive sera, 197 were positive for both HCV Ag (≥3 fmol/L) and HCV RNA (>15 IU/mL), 22 were negative for both tests, while 2 were positive to HCV RNA only. The sensitivity and specificity for HCV Ag in predicting HCV RNA were 99% and 100%, respectively. Out of 199 patients (90%) tested positive for HCV viremia, 107 (56%) were of genotype 1, 77 (38.7%) of genotype 2 and 15 of other genotypes. Analysis of 221 anti-HCV-positive patient sera found a strong positive correlation between HCV RNA and HCV-Ag (r = 0.960, p < 0.001). Genotype 1 (log [HCV RNA] = 0.988 x log [HCV-Ag] + 2.768), with correlation coefficient 0.945, exhibited a stronger correlation than genotype 2 (log [HCV RNA] = 0.859 x log [HCV-Ag] + 2.859; r = 0.862). Given the strong positive correlation between HCV-Ag immunoassay and HCV RNA molecular assay in genotyping affected individuals, we propose that HCV-Ag immunoassay is a more cost and time efficient alternative to the current two-step diagnostic process.

A voltammetric immunoassay for the carcinoembryonic antigen using a self-assembled magnetic nanocomposite.

The authors describe a voltammetric immunoassay for the carcinoembryonic antigen (CEA). It is based on the use of a self-assembled magnetic nanocomposite as multifunctional signal amplification platform. The core of the nanocomposite consists of Fe3O4 microspheres, and the shell of zirconium hexacyanoferrate loaded with gold nanoparticles (AuNPs@ZrHCF@Fe3O4). The material was synthesized by an electrostatic self-assembly process which is caused by the strong interaction between cyano groups and AuNPs. The surface of the Fe3O4 microspheres was functionalized with amino groups to facilitate the immobilization of ZrHCF which acts as an electron mediator. The nanocomposite was placed on a glassy carbon electrode which then displays noteworthy electrocatalytic activity toward the reduction of hydrogen peroxide (H2O2). The AuNPs serve as a support for the immobilization of antibodies by the interaction between AuNPs and amino groups on antibodies to construct a covalent Au-N bond. This facilitates electron transfer on the electrode surface using H2O2 as the electrochemical probe. Square wave voltammetry (measured typically at +0.2V vs. SCE) was carried out to record the electrochemical behavior. Under the optimal conditions, a response is linear in the 0.5pg·mL-1 to 50ng·mL-1 CEA concentration range, and the detection limit is as low as 0.15pg·mL-1 (S/N= 3). The method is selective, highly stable and acceptably reproducible. Graphical abstract A self-assembly magnetic nanocomposite for voltammetric immunoassay of CEA. GCE glassy carbon electrode; Au NPs gold nanoparticles; ZrHCF zirconium hexacyanoferrate; CEA carcinoembryonic antigen; Anti-CEA CEA antibody; BSA bovine serum albumin; SWV square wave voltammetry. A high sensitive voltammetric immunoassay method has been used for detecting CEA, It is based on a self-assembled magnetic nanocomposite (Au NPs@ZrHCF@Fe3O4) as multifunctional signal amplification platform.

Detection of Influenza A and B Viruses and Respiratory Syncytial Virus by Use of Clinical Laboratory Improvement Amendments of 1988 (CLIA)-Waived Point-of-Care Assays: a Paradigm Shift to Molecular Tests

An accurate laboratory diagnosis of influenza, respiratory syncytial virus (RSV), and other respiratory viruses can help to guide patient management, antiviral therapy, infection prevention strategies, and epidemiologic monitoring. Influenza has been the primary driver of rapid laboratory testing due to its morbidity and mortality across all ages, the availability of antiviral therapy, which must be given early to have an effect, and the constant threat of new pandemic strains. Over the past 30 years, there has been an evolution in viral diagnostic testing, from viral culture to rapid antigen detection, and more recently, to highly sensitive nucleic acid amplification tests (NAAT), as well as a trend to testing at the point of care (POC). Simple rapid antigen immunoassays have long been the mainstay for POC testing for influenza A and B viruses and respiratory syncytial virus (RSV) but have been faulted for low sensitivity. In 2015, the first POC NAAT for the detection of influenza was approved by the Food and Drug Administration (FDA), ushering in a new era. In 2017, the FDA reclassified rapid influenza diagnostic tests (RIDTs) from class I to class II devices with new minimum performance standards and a requirement for annual reactivity testing. Consequently, many previously available RIDTs can no longer be purchased in the United States. In this review, recent developments in Clinical Laboratory Improvement Amendments of 1988 (CLIA)-waived testing for respiratory virus infections will be presented, with the focus on currently available FDA-cleared rapid antigen and molecular tests primarily for influenza A and B viruses and RSV.

Dye sensitized photoelectrochemical immunosensor for the tumor marker CEA by using a flower-like 3D architecture prepared from graphene oxide and MoS2.

The authors describe a dye-sensitized photoelectrochemical immunoassay for the tumor marker carcinoembryonic antigen (CEA). The method employs the rhodamine dye Rh123 with red color and absorption maximum at 500nm for spectral sensitization, and a 3D nanocomposite prepared from graphene oxide and MoS2 acting as the photoelectric conversion layer. The nanocomposite with flower-like 3D architectures was characterized by transmission electron microscopy, scanning electron microscopy, X-ray powder diffraction, and UV-vis diffuse reflectometry. A photoelectrochemical sandwich immunoassay was developed that is based on the use of the nanocomposite and based on the specific binding of antibody and antigen, and by using a secondary antibody labeled with Rh123 and CdS (Ab2-Rh123@CdS). Under optimal conditions and at a typical working voltage of 0V (vs. Hg/HgCl2), the photocurrent increases linearly 10pgmL-1 to 80ngmL-1 CEA concentration range, with a 3.2pgmL-1 detection limit. Graphical abstract Flower-like GO-MoS2 complex with high efficiency of electron transport was synthesized to construct photoelectrochemical platform. The sandwich-type immunoassay was built on this platform based on specific binding of antigen and antibody. Carcinoembryonic antigen in sample was detected sensitively by using sensitization of rhodamine dye Rh123 as signal amplification strategy.

Electrochemical immunoassay for the carcinoembryonic antigen based on the use of a glassy carbon electrode modified with an octahedral Cu2O-gold nanocomposite and staphylococcal protein for signal amplification.

The authors describe an electrochemical immunoassay for ultrasensitive direct determination of the carcinoembryonic antigen (CEA). A nanocomposite consisting of octahedral Cu2O nanocrystals covered with gold nanoparticles was utilized to modify a glassy carbon electrode which gives a strongly enhanced chronoamperometric signal for H2O2 which is used as an electrochemical probe. The morphology and elemental composition of the the nanocomposite was studied by field emission scanning electron microscopy and energy dispersive X-ray spectroscopy. In addition, staphylococcal protein A was placed on the electrode for efficient capture of antibody to further enhance the sensitivity of the assay. Under optimal conditions and at a typical working voltage of -0.4V (vs. Ag/AgCl), the response covers the 2pg·mL-1 to 20ng·mL-1 CEA concentration range with a 200fg·mL-1 lower detection limit. The method was successfully applied to the determination of CEA in (spiked) human serum. Graphical abstract Schematic of the fabrication of an electrochemical immunosensor for ultrasensitive detection the carcinoembryonic antigen. The sensor is based on the use of a glassy carbon electrode modified with an octahedral Cu2O-gold nanocomposite and staphylococcal protein A for signal amplification.

Field validation of recombinant antigen immunoassays for diagnosis of Lassa fever

Lassa fever, a hemorrhagic fever caused by Lassa virus (LASV), is endemic in West Africa. It is difficult to distinguish febrile illnesses that are common in West Africa from Lassa fever based solely on a patient’s clinical presentation. The field performance of recombinant antigen-based Lassa fever immunoassays was compared to that of quantitative polymerase chain assays (qPCRs) using samples from subjects meeting the case definition of Lassa fever presenting to Kenema Government Hospital in Sierra Leone. The recombinant Lassa virus (ReLASV) enzyme-linked immunosorbant assay (ELISA) for detection of viral antigen in blood performed with 95% sensitivity and 97% specificity using a diagnostic standard that combined results of the immunoassays and qPCR. The ReLASV rapid diagnostic test (RDT), a lateral flow immunoassay based on paired monoclonal antibodies to the Josiah strain of LASV (lineage IV), performed with 90% sensitivity and 100% specificity. ReLASV immunoassays performed better than the most robust qPCR currently available, which had 82% sensitivity and 95% specificity. The performance characteristics of recombinant antigen-based Lassa virus immunoassays indicate that they can aid in the diagnosis of LASV Infection and inform the clinical management of Lassa fever patients.

A novel electrochemical immunoassay for carcinoembryonic antigen based on glucose oxidase-encapsulated nanogold hollow spheres with a pH meter readout.

A portable electrochemical immunosensing protocol was designed for the sensitive detection of a disease-related tumor biomarker (carcinoembryonic antigen, CEA, used in this case) on a pH meter using glucose oxidase (GOx)-encapsulated gold hollow microspheres (AuHMs) for signal amplification. The assay was carried out on a monoclonal anti-CEA capture antibody-coated microplate with a sandwich-type reaction mode. The GOx-entrapped AuHM was first synthesized using the reverse micelle method and then used as the signal-generation tag for the labeling of polyclonal anti-CEA detection antibody. Accompanying the formation of the sandwiched immunocomplexes, the loaded GOx molecules in the microsphere could catalyze glucose into gluconic acid and hydrogen peroxide. The as-produced gluconic acid changed the microenvironment of the detection solution, thus resulting in the shift of the pH value, which could be quantitatively determined on a portable pH meter. The use of gold hollow microspheres was expected to enhance the loaded amount of GOx for signal amplification. Two labeling protocols including GOx-labeled secondary antibody and GOx-AuHM-labeled secondary antibody were investigated for CEA detection, and improved analytical features were acquired with GOx-AuHM labeling. With the GOx-AuHM labeling strategy, the pH meter-based immunosensing device exhibited a good analytical performance for CEA detection within the dynamic linear range of 0.1-100 ng mL-1 at a detection limit of 0.062 ng mL-1. The strong attachment of anti-CEA antibodies to GOx-AuHM brought a good repeatability and intermediate precision down to 10%. Importantly, no significant differences at the 0.05 significance level were encountered in the analysis of 12 human serum specimens between the developed immunoassay and the commercialized electrochemiluminescent method for CEA determination.