Antigen Assay Research Articles

Integrated Antigenic and Nucleic Acid Detection in Single Virions and Extracellular Vesicles with Viral Content.

Virion-mediated outbreaks are imminent and despite rapid responses, continue to cause adverse symptoms and death. Therefore, tunable, sensitive, high-throughput assays are needed to help diagnose future virion-mediated outbreaks. Herein, it is developed a tunable in situ assay to selectively enrich virions and extracellular vesicles (EVs) and simultaneously detect antigens and nucleic acids at a single-particle resolution. The Biochip Antigen and RNA Assay (BARA) enhanced sensitivities compared to quantitative reverse-transcription polymerase chain reaction (qRT-PCR), enabling the detection of virions in asymptomatic patients, genetic mutations in single virions, and enabling the continued long-term expression of viral RNA in the EV-enriched subpopulation in the plasma of patients with post-acute sequelae of the coronavirus disease of 2019 (COVID-19). BARA revealed highly accurate diagnoses of COVID-19 by simultaneously detecting the spike glycoprotein and nucleocapsid-encoding RNA in saliva and nasopharyngeal swab samples. Altogether, the single-particle detection of antigens and viral RNA provides a tunable framework for the diagnosis, monitoring, and mutation screening of current and future outbreaks.

Synergistic effects of glial cell line-derived neurotrophic factor and base-medium on in vitro culture of testicular tissue derived from prepubertal collared peccary.

We evaluated the influence of different media plus various concentrations of Glial cell line-derived neurotrophic factor (GDNF) during the in vitro culture (IVC) of testicular tissues from prepubertal collared peccary. Testes from 5 individuals were collected, fragmented and cultured for 28 days (34°C and 5% CO2). Culture media were Dulbecco's modified essential medium (DMEM) or stem cell serum free media (StemPro-34™ SFM), both supplemented with various concentrations of GDNF (0, 10, or 20 ng/mL). Fragments were cultured on the flat surface of 0.75% agarose gel and were evaluated every 7 days for fragment area, histomorphology, cellular viability, and proliferative activity. Data were expressed as mean ± standard error and analyzed by Kruskal-Wallis's and Tukey test. Fragments area decreased over the 28 days-culture, regardless of the treatment. For morphology, the StemPro-37 SFM medium plus 10 ng/mL GDNF provided higher scores at all time points in comparison to DMEM using any GDNF concentration (p < .05). After 28 days, similar cellular viability (~70%) was observed in all treatments (p > .05). For proliferating cell nuclear antigen assay, only DMEM plus 10 ng/mL GDNF improved (p < .05) cellular proliferation on Days 14 and 28. Looking at argyrophilic nucleolar organizing regions, after 28 days, there were no differences among treatments regarding cell proliferative capacity for both spermatogonia and Sertoli cells (p > .05). In summary, the DMEM and StemPro-34 SFM are adequate medium for IVC of prepubertal peccary testicular tissue. Supplementation with GDNF, especially at a 10 ng/mL concentration, appears to be essential for the maintenance of cell survival and proliferation.

Case - Biotin supplements interfering with prostate-specific antigen assays A cautionary tale.

Case - Biotin supplements interfering with prostate-specific antigen assays A cautionary tale.

Monitoring Influenza C and D Viruses in Patients With Respiratory Diseases in Japan, January 2018 to March 2023.

Influenza viruses can cause zoonotic infections that pose public health risks. Surveillance of influenza A and B viruses is conducted globally; however, information on influenza C and D viruses is limited. Longitudinal monitoring of influenza C virus in humans has been conducted in several countries, but there has been no long-term monitoring of influenza D virus in humans. The public health risks associated with the influenza D virus therefore remain unknown. We established a duplex real-time RT-PCR to detect influenza C and D viruses and analyzed respiratory specimens collected from 2144 patients in Japan with respiratory diseases between January 2018 and March 2023. We isolated viruses and conducted hemagglutination inhibition tests to examine antigenicity and focus reduction assays to determine susceptibility to the cap-dependent endonuclease inhibitor baloxavir marboxil. We detected three influenza C viruses belonging to the C/Kanagawa- or C/Sao Paulo-lineages, which recently circulated globally. None of the specimens was positive for the influenza D virus. The C/Yokohama/1/2022 strain, isolated from the specimen with the highest viral RNA load and belonging to the C/Kanagawa-lineage, showed similar antigenicity to the reference C/Kanagawa-lineage strain and was susceptible to baloxavir. Our duplex real-time RT-PCR is useful for the simultaneous detection of influenza C and D viruses from the same specimen. Adding the influenza D virus to the monitoring of the influenza C virus would help in assessing the public health risks posed by this virus.

An Analogy between Gold Standard SARS-CoV-2 RT-qPCR with the SARS-CoV-2 Rapid Antigen Test in a Tertiary Care Setting in Central State of India

Reverse transcription-quantitative PCR (RT-qPCR)-based assays are extensively being utilized to detect coronavirus disease 2019 (COVID-19). However, due to a lack of RT-qPCR testing capability, these tests cannot be carried out in community clinics. The intention of our study was to evaluate the specificity and sensitivity of Rapid Antigen Detection (RAT) tests versus those of RT-qPCR using nasopharyngeal and oropharyngeal specimens. Respiratory swab specimens were collected from the COVID-19 patients admitted at Dr. Bhimrao Ambedkar Memorial Hospital, Raipur, CG, India, during March to April 2022. RAT and RT-qPCR were performed using standard methods as per guidebook instructions, and subjects were chosen using a convenience sample technique. 100 swabs from patients, who had earlier verified positive and 100 from who had earlier verified negative for SARS-CoV-2 via RT-qPCR, were taken for study. Study was approved by the institutional ethical committee before data collection and initiation of the study. We evaluated for the sensitivity and specificity of the STANDARD Q COVID-19 Ag test kit (SD Biosensor). On testing, an over-all sensitivity and specificity of the kit was recorded as 74% and 100%, respectively in comparison to the RT-qPCR kit. Further, the assay’s sensitivity was shown to be 100%, 94.87%, 77.27%, and 55.56%, respectively, for samples with cycle thresholds (Ct) of 15-25, 25-30, 30-35, and &gt;35. We draw the conclusion that the RT-qPCR assay has superior sensitivity and specificity to the antigen assay. However, in all situations where RT-qPCR testing is difficult, the antigen assay could serve as a rapid and simple option for separating SARS-CoV-2 contagious from non-contagious patients.

Thrombophilia Screening: Not So Straightforward.

Although inherited thrombophilias are lifelong risk factors for a first thrombotic episode, progression to thrombosis is multifactorial and not all individuals with inherited thrombophilia develop thrombosis in their lifetimes. Consequently, indiscriminate screening in patients with idiopathic thrombosis is not recommended, since presence of a thrombophilia does not necessarily predict recurrence or influence management, and testing should be selective. It follows that a decision to undertake laboratory detection of thrombophilia should be aligned with a concerted effort to identify any significant abnormalities, because it will inform patient management. Deficiencies of antithrombin and protein C are rare and usually determined using phenotypic assays assessing biological activities, whereas protein S deficiency (also rare) is commonly detected with antigenic assays for the free form of protein S since available activity assays are considered to lack specificity. In each case, no single phenotypic assay is capable of detecting every deficiency, because the various mutations express different molecular characteristics, rendering thrombophilia screening repertoires employing one assay per potential deficiency, of limited effectiveness. Activated protein C resistance (APCR) is more common than discrete deficiencies of antithrombin, protein C, and protein S and also often detected initially with phenotypic assays; however, some centres perform only genetic analysis for factor V Leiden, as this is responsible for most cases of hereditary APCR, accepting that acquired APCR and rare F5 mutations conferring APCR will go undetected if only factor V Leiden is evaluated. All phenotypic assays have interferences and limitations, which must be factored into decisions about if, and when, to test, and be given consideration in the laboratory during assay performance and interpretation. This review looks in detail at performance and limitations of routine phenotypic thrombophilia assays.

Rare bleeding disorders in Egyptian females presented with heavy menstrual bleeding: single-center study

BackgroundHeavy menstrual bleeding is an important health problem in women of reproductive age and is also one of the most common symptoms in women with bleeding disorders. Data about the frequency of rare bleeding disorders are limited, and population-based studies are lacking, so we aimed to determine the frequency of rare bleeding disorders among women presented with heavy menstrual periods that cannot be attributed to obvious problems.MethodsComplete blood count and bleeding profiles include activated partial thromboplastin time, prothrombin time, factor VIII activity assay, ristocetin cofactor activity, von Willebrand antigen assay, platelet aggregation tests and other factor assays in 100 out of 300 females presented with unexplained heavy menstrual period, pictorial bleeding assessment chart (PBAC) > 100 as a screening tool for heavy menstrual periods, and or International Society of Thrombosis and Hemostasis-Bleeding Assessment Tool (ISTH-BAT) > 6.ResultsA total of 300 women with heavy menstrual periods without an obvious explained cause were included in our study. Among them, we found 100 (30%) females with a mean PBAC of 234 ± 147 and mean ISTH-BAT of 9 ± 5 denoting HMB may be due to underlying bleeding disorders. Among them, the most common diagnosis was VWD in 30 (30%). Other disorders were as follows: 28 (28%) cases with clotting factor deficiencies, 24 (24%) cases were found to have platelet dysfunction, and in 18% of our studied cases, we did not find a clear cause of their bleeding disorders (unknown).ConclusionRare bleeding disorders are not uncommon and require comprehensive hemostatic evaluation as well as simple tools like PBAC and ISTH-BAT questionnaires for the identification of females presented with unexplained HMB.

Diagnostic accuracy of a novel lateral flow assay for histoplasmosis.

Antigen testing is an important diagnostic tool for histoplasmosis but has limited availability globally. We evaluated the OIDx urine lateral flow antigen assay among 204 persons suspected to have histoplasmosis. Among patients with proven histoplasmosis, sensitivity was 33.3% (3/9, 95% CI 7.5%-70.1%) and specificity 80.5% (157/195, 95% CI 74.3%-85.8%). The MiraVista urine antigen test had better specificity (96.9%) and equal sensitivity. The OIDx test demonstrated 33.3% (3/9) positive agreement and 84.0% (163/194) negative agreement with the MiraVista test. These results should be considered in the context of our low HIV prevalence population with a mixture of pulmonary and disseminated disease.

Analytical performance evaluation of breast and ovarian cancer antigen assays on the Atellica CI analyzer

Analytical performance evaluation of breast and ovarian cancer antigen assays on the Atellica CI analyzer

Performance evaluation of the architect hepatitis c core antigen assay vs roche HCV DUO

Performance evaluation of the architect hepatitis c core antigen assay vs roche HCV DUO

Hematological profile of pediatric patients with dengue in district hospital Samba

Background: Dengue is an acute viral illness spread by vector aedes mosquito and caused by RNA virus of the family Flaviviridae. Dengue presents with high fever, severe headache, vomiting, arthralgia, myalgia. Laboratory findings include thrombocytopenia and leukopenia. Dengue is self-limiting illness. Objectives were to study hematological profile of pediatric patients with dengue in district hospital samba from September to November 2022. Methods: Dengue patients aged 0-18 years detected by ELISA NS1 antigen assay in DH samba were included in the study. CBC of all these patients was analyzed on day of presentation to hospital and day of dengue positivity. Results: There were 50 participants in this study. Out of these 35 (70%) were males and 15 (30%) were females, 11 patients were 1-5-year-old (22%); 16 patients were 6-12-year-old (32%); 23 patients were 13-18-year-old (46%), 21 patients (42%) had thrombocytopenia at presentation. 29 out of 50 (58%) were anemic, 19 out of 50 (38%) patients had leukocytopenia. Statistically significant association were seen between age group and platelet counts; platelet and leukocyte count. Conclusions: Teenagers were most commonly affected. Males were affected 3× more than females. Although thrombocytopenia was not universal at presentation but it was observed in less than half (42%) of study participants. One third of study participants (38%) had leukocytopenia at presentations. Thrombocytopenia was most predominant in adolescent age group. Leukocytopenia and thrombocytopenia were associated with each other.

P150 Diagnostic utility of anti-cN1A autoantibody testing in sporadic inclusion body myositis

Abstract Background/Aims Sporadic inclusion body myositis (sIBM) is the most common acquired myopathy in patients aged over 50 years. Diagnostic criteria encompass clinical, biochemical, and myopathological features. Certain myopathological features possess high specificity but low sensitivity; reliance on such findings may lead to delayed or misdiagnoses. The discovery of autoantibodies against cytosolic 5'-Nucleotidase 1A (cN1A) in sIBM patients suggests an autoimmune component in pathogenesis, but the inclusion of this autoantibody in diagnostic criteria remains uncertain. Additionally, optimal testing methodology has not been established, with varying techniques (e.g., ELISA/line-blot) in use. We assessed antibody profiles in an sIBM cohort and reviewed diagnoses in anti-cN1A positive patients. Methods This study was based at the Northern Care Alliance NHS Foundation Trust which hosts a tertiary neuromuscular referral service for the North-West of England. Data sources included myositis clinic and immunology results databases. All patients tested using a validated myositis line-blot immunoassay (EUROLINE 16 Antigen et cN1A [EUROIMMUN]) November 2021-April 2023 were included. Diagnoses of all tested patients were identified through case-note review. Sensitivity and specificity metrics were calculated based on diagnosis and anti-cN1A antibody status. Results The total number of patients tested for anti-cN1A was 495. Of these, 10 patients had a diagnosis of sIBM. The true-positive and false-negative rates were both 50% (n = 5 each). Of the 485 patients who did not have sIBM, true-negativity was 97.52% (n = 473) and false-positivity 2.47% (n = 12). Overall, anti-cN1A had a sensitivity of 50%, specificity of 97.52%, positive predictive value of 29.41% and negative predictive value of 98.95% for sIBM (Table 1). Patients with a false-positive anti-cN1A antibody had diagnoses of: primary Raynaud’s (n = 5); undifferentiated CTD (n = 3); systemic sclerosis (n = 2); non-sIBM myositis (n = 2). Conclusion Anti-cN1A detected using the EUROLINE 16 Antigen assay has high specificity but lacks sensitivity for sIBM, consistent with previous literature. These findings suggest that this assay performs similarly to other cN1A detection methodologies and has utility in confirming the diagnosis of sIBM where there is clinical suspicion. Further real-world analysis of antibody data in larger cohorts is required to clarify the role of serological testing in sIBM diagnostic criteria, to improve diagnostic delays and misclassifications. Disclosure M. Al-Attar: None. T. Khoo: None. J.B. Lilleker: None. H. Chinoy: None.

Is there an interest in systematic serum screening for aspergillosis in COVID-19 patients in a medical ward?

PurposeWe evaluated the interest of systematic screening of serum fungal markers in patients hospitalized in a medical ward. MethodsWe retrospectively analyzed all patients hospitalized in our infectious disease department from October 1st to October 31st, 2020 for COVID-19 without prior ICU admission, and for whom systematic screening of serum fungal markers was performed. ResultsThirty patients were included. The majority of patients received corticosteroids (96.7%). The galactomannan antigen assay was positive for 1/30 patients at D0, and 0/24, 0/16, 0/13 and 0/2 at D4, D7, D10 and D14 respectively. 1,3-ß-D-glucan was positive for 0/30, 1/24, 1/12, 0/12, 0/2 at D0, D4, D7, D10 and D14 respectively. No Aspergillus fumigatus PCR was positive. No cases of aspergillosis were retained. ConclusionOur study does not support the interest of systematic screening of fungal markers in immunocompetent patients with COVID-19 in a conventional unit.

Hepatitis C virus core antigen: A diagnostic and treatmentmonitoring marker of hepatitis C virus in Indian population.

The diagnosis and treatment monitoring of hepatitis C is quite challenging. The screening test, i.e. antibody assay, is unable to detect acute cases, while the gold standard hepatitis C virus (HCV) reverse transcriptase polymerase chain reaction (RTPCR) assay is not feasible in resource-limited countries such as India due to high cost and infrastructure requirement. European Association for the Study of the Liver and World Health Organization have approved a new marker, i.e. HCV core antigen (HCVcAg) assay, as an alternative to molecular assay. In this study, we have evaluated HCVcAg assay for diagnosis and treatment monitoring follow-up in Indian population infected with hepatitis C. Blood specimen of 90 clinically suspected cases of acute hepatitis C were tested simultaneously for anti-HCV antibody assay via ELISA (enzyme-linked immunoassay), HCVcAg assay by chemiluminescence immune assay (CLIA) and HCV RTPCR VL (viral load) assay. Thirty-four HCV RTPCR positive patients were further enrolled in treatment monitoring group whose blood samples were tested at thebeginning of treatment, twoweeks, fourweeks and 12weeks via HCV core Ag assay and HCV RTPCR Viral Load assay. Considering HCV RTPCR as gold standard, diagnostic performance of HCV core Ag assay and anti-HCV antibody assay was evaluated. The sensitivity and specificity of HCV core Ag assay were higher than that of anti-HCV Antibody assay, i.e. 88.3% and 100% vs. 23.3% and 83.3%, respectively. The overall diagnostic accuracy of HCV core Ag assay was 92.20%. Among treatment follow-up group, HCV core Ag levels correlated well with HCV viral load levels, at the beginning of treatment (baseline) till 12weeks showing highly significant Spearman rank correlation coefficient of > 0.9 with HCV viral load levels. HCV core Ag assay is a cost-effective, practically feasible substitute of HCV RTPCR viral load assay for diagnosis as well as long duration treatment monitoring of hepatitis C infection in resource-limited settings.

Platelets retain function and can be stored following disruption of human leucocyte antigens.

Antibodies to human leucocyte antigen (HLA) Class-I antigens can lead to refractoriness to platelet transfusion. Although this can be overcome by transfusion of HLA-compatible platelets, they are not always available. Disruption of HLA antigens on platelets by acid treatment may be a suitable alternative when no other components are available. The aim of this study was to assess the effect of HLA disruption and subsequent storage of platelet components. Platelet components were treated with 0.9% saline or citric acid solution (pH 3.0), and then stored until expiry (Day 7). HLA and platelet glycoprotein expression, platelet viability, activation and sialylation were measured by flow cytometry. Release of soluble factors was measured by ELISA and metabolism by biochemistry analyser. Reactivity to patient anti-sera containing anti-HLA antibodies was measured using platelet immunofluorescence tests (PIFTs) and monoclonal antibody immobilization of platelet antigen (MAIPA) assays. Platelet function was measured using aggregometry and thromboelastography (TEG). Acid treatment reduced detection of HLA Class-I on platelets by 75%, with significant reductions in reactivity to patient anti-sera. Acid treatment reduced platelet content and viability, increased platelet activation and accelerated metabolism. Glycan cleavage was increased by acid treatment. Treatment reduced platelet activation following agonist stimulation by ADP and TRAP-6, but platelets remained functional, displaying increased aggregation response and reduced time to clot formation by TEG. Although HLA disruption had some detrimental effects, acid-treated platelets remained functional, retaining their capacity to respond to agonists and form clots, and with further development could be used to support refractory patients.

Galactomannan Antigen Assay for the Diagnosis of Invasive Aspergillosis: A Study from a Tertiary Care Hospital of Central India

Invasive Aspergillosis (IA) is a severe and fatal infection, especially in immunocompromised patients. Galactomannan is a polysaccharide antigen present in the cell wall of Aspergillus species, which is secreted into the blood and other body fluids during hyphal growth. Therefore, detecting galactomannan antigen is very useful in diagnosing IA, along with clinical features and radiological findings. The study period was one year (January 2022 to December 2022). The data was collected retrospectively from the medical records and case sheets of all clinically suspected invasive aspergillosis patients. Galactomannan antigen assay was performed using an FDA-approved Platelia Aspergillus EIA test kit, and results were interpreted according to the manufacturer’s instructions (Cut off &gt; 0.5). A total of 236 clinically suspected Invasive Aspergillosis cases were enrolled in the study. Galactomannan positivity was predominantly seen in patients aged 40 – 60 years, with male preponderance. Of 236 patients, 14.40% were immunocompromised, and 85.59% were immunocompetent. According to EORTC/MSG definitions, we got one proven IA case, 21 probable cases (8.89%), and nine (3.81%) possible cases. In immunocompetent individuals also, we observed 48.72% (115/236) galactomannan positivity, especially in old Pulmonary Tuberculosis (PTB) patients. Galactomannan positivity was higher in Broncho alveolar lavage (BAL) samples (n=70, 85.36%) than in serum samples (n=77, 46.67%). We found culture positivity of 14.06%, with Aspergillus fumigatus being the commonest isolate, followed by Aspergillus flavus. There is increased positivity of galactomannan in BAL samples compared to serum; hence BAL is a better specimen for diagnosis of Invasive Pulmonary Aspergillosis (IPA).

Diagnosis of thrombotic thrombocytopenic purpura: easy-to-use fiber optic surface plasmon resonance immunoassays for automated ADAMTS-13 antigen and conformation evaluation

BackgroundLaboratory diagnosis of immune-mediated thrombotic thrombocytopenic purpura (iTTP) remains challenging when ADAMTS-13 activity ranges between 10% and 20%. To prevent misdiagnosis, open ADAMTS-13 conformation gained clinical attention as a novel biomarker, especially to diagnose acute iTTP in patients with diagnostic undecisive ADAMTS-13 activity. Plasma ADAMTS-13 conformation analysis corrects for ADAMTS-13 antigen, with both parameters being characterized in enzyme-linked immunosorbent assay (ELISA)-based reference assays requiring expert technicians. ObjectivesTo design ADAMTS-13 antigen and conformation assays on automated, easy-to-use fiber optic surface plasmon resonance (FO-SPR) technology to promote assay accessibility and diagnose challenging iTTP patients. MethodsADAMTS-13 antigen and conformation assays were designed on FO-SPR technology. Plasma of 20 healthy donors and 20 acute iTTP patients were quantified, and data from FO-SPR and ELISA reference assays were compared. ResultsFollowing assay design, both antigen and conformation FO-SPR assays were optimized and characterized, presenting strong analytical sensitivity (detection limit of 0.001 μg/mL) and repeatability (interassay variation of 14.4%). Comparative analysis suggested positive correlation (Spearman r of 0.92) and good agreement between FO-SPR and ELISA assays. As expected, FO-SPR assays showed a closed or open ADAMTS-13 conformation in healthy donors and acute iTTP patients, respectively. ConclusionBoth ADAMTS-13 antigen and conformation assays were transferred onto automated, easy-to-use FO-SPR technology, displaying potent analytical sensitivity and reproducibility. ADAMTS-13 antigen and conformation were determined for healthy donors and acute iTTP patients showing strong correlation with ELISA reference. Introducing FO-SPR technology in clinical context could support routine diagnosis of acute iTTP patients, notably when ADAMTS-13 activity fluctuates between 10% and 20%.

Predictive Value of Transrectal Ultrasonic Doppler and Elastographic Features in Prostate Cancer Detection in Lagos University Teaching Hospital.

This study aimed at determining the predictive value (PV) of transrectal ultrasonic Doppler and elastographic features in prostate cancer (PCa) detection among patients in Lagos University Teaching Hospital. This prospective study involved patients that underwent evaluation for PCa. Participants had digital rectal examination (DRE), prostate-specific antigen (PSA) assay, and transrectal ultrasound-guided prostate biopsy using colour Doppler (CD) and elastography. All cores were sent for histopathology. Data were analysed using Statistical Package for the Social Sciences Version 22.0. CD and elastography PV in PCa detection and their relationships to the Gleason score (GS) were analysed (P < 0.05). Seventy men (aged between 45 and 87 years) were enrolled. Forty-three (61.4%) patients had PCa with a mean age of 69.37 ± 8.22years. The sensitivity, specificity, positive PV (PPV), negative PV (NPV) and accuracy of CD were 8.50%, 97.44%, 64.10%, 66.42% and 66.31%, respectively. The sensitivity, specificity, PPV, NPV and accuracy of elastography were 84.21%, 94.59%, 88.89%, 92.11% and 91.07%, respectively. There is a significant association between decreased elasticity (elastography) and PCa detection but a weak association between increased vascularity (CD) and PCa detection. A positive correlation exists between extent of prostatic stiffness and GS.

Comparative Diagnostic Value of Dengue Infection Using ELFA and Two Commercial Immuno-Chromatography Tests

Dengue Virus (DEN-V) can lead to a broad spectrum of clinical presentations, from mild symptoms to mortality. Based on the presence of antibodies, dengue infection is categorized into primary and secondary dengue. Early diagnosis is crucial for effective treatment of DEN-V infection. Non-Structural Antigen 1 (NS1) and dengue antibodies like immunoglobulin M (IgM) and immunoglobulin G (IgG) anti-dengue are standard assays used for dengue diagnosis. Various immunoassay methods, including Enzyme-Linked Fluorescent Assay (ELFA) and Immuno-Chromatographic Tests (ICT), are employed to detect these antigens and antibodies. This study aimed to compare the diagnostic value between ELFA and two commercial ICTs for detecting NS1 antigen, IgM/IgG anti-dengue. Seventy suspected dengue patients with fever lasting two to seven days at Premier Hospital, Surabaya were enrolled in this study. Blood serum samples from the individuals were tested for NS1 antigen and IgM/IgG anti-dengue using VIDAS® and two commercial ICTs (Boson and SD Bioline). The sensitivity of NS1 antigen assays using ELFA and two ICTs was calculated, as well as the agreement rate between ELFA and both ICTs. ELFA demonstrated high sensitivity (77.97%) for NS1 antigen detection in diagnosing dengue infection compared to two ICTs (76.27% and 45.45%). Using an automated system in ELFA can offer more excellent diagnostic value and objective results and determine the cut-off ratio of IgM/IgG antibodies. All of this comes at a cost comparable to ICT.

A label-free aptasensing method for detecting SARS-CoV-2 virus antigen by using dumbbell probe-mediated circle-to-circle amplification.

Controlling the spread of pathogen requires an efficient and accurate diagnosis. Compared with nucleic acid and antibody detection, antigen assays are more convenient to meet clinical diagnostic needs. However, antigen detection is often difficult to achieve high sensitivity in a limited time. In this work, a novel aptasensing method was designed for the purpose of SARS-CoV-2 antigen detection, using a dumbbell padlock probe-mediated circle-to-circle amplification (C2CA) approach. A sandwich complex of antibody-antigen-aptamer is first formed on the magnetic beads. Afterwards, the signal is amplified by a C2CA reaction involving two tandem rolling circle amplifications. Without special instruments or nanomaterials, a detection limit of 575fg/mL for S1 protein can be achieved in less than 2h. In the case of the spike pseudovirus SARS-CoV-2 in artificial saliva, the detection limit is 272 TU/μL, which is much lower than average viral load in patients. Therefore, our method provides a timely, efficient and accurate approach for the clinical diagnosis of SARS-CoV-2. It also opens up the application of C2CA in aptamer sensing and antigen detection.