Next-Generation Multitarget Stool DNA Test for Colorectal Cancer Screening.

BACKGROUND: A next-generation multitarget stool DNA test, including assessments of DNA molecular markers and hemoglobin level, was developed to improve the performance of colorectal cancer screening, primarily with regard to specificity. METHODS: In a prospective study, we evaluated a next-generation multitarget stool DNA test in asymptomatic adults 40 years of age or older who were undergoing screening colonoscopy. The primary outcomes were sensitivity of the test for colorectal cancer and specificity for advanced neoplasia (colorectal cancer or advanced precancerous lesions). Advanced precancerous lesions included one or more adenomas or sessile serrated lesions measuring at least 1 cm in the longest dimension, lesions with villous histologic features, and high-grade dysplasia. Secondary objectives included the quantification of sensitivity for advanced precancerous lesions and specificity for nonneoplastic findings or negative colonoscopy and comparison of sensitivities for colorectal cancer and advanced precancerous lesions between the multitarget stool DNA test and a commercially available fecal immunochemical test (FIT). RESULTS: Of 20,176 participants, 98 had colorectal cancer, 2144 had advanced precancerous lesions, 6973 had nonadvanced adenomas, and 10,961 had nonneoplastic findings or negative colonoscopy. With the next-generation test, sensitivity for colorectal cancer was 93.9% (95% confidence interval [CI], 87.1 to 97.7), and specificity for advanced neoplasia was 90.6% (95% CI, 90.1 to 91.0). Sensitivity for advanced precancerous lesions was 43.4% (95% CI, 41.3 to 45.6), and specificity for nonneoplastic findings or negative colonoscopy was 92.7% (95% CI, 92.2 to 93.1). With the FIT, sensitivity was 67.3% (95% CI, 57.1 to 76.5) for colorectal cancer and 23.3% (95% CI, 21.5 to 25.2) for advanced precancerous lesions; specificity was 94.8% (95% CI, 94.4 to 95.1) for advanced neoplasia and 95.7% (95% CI, 95.3 to 96.1) for nonneoplastic findings or negative colonoscopy. As compared with FIT, the next-generation test had superior sensitivity for colorectal cancer (P<0.001) and for advanced precancerous lesions (P<0.001) but had lower specificity for advanced neoplasia (P<0.001). No adverse events occurred. CONCLUSIONS: The next-generation multitarget stool DNA test showed higher sensitivity for colorectal cancer and advanced precancerous lesions than FIT but also showed lower specificity. (Funded by Exact Sciences; BLUE-C ClinicalTrials.gov number, NCT04144738.).

Testing for SARS-CoV-2: lessons learned and current use cases.

SUMMARYThe emergence and worldwide dissemination of SARS-CoV-2 required both urgent development of new diagnostic tests and expansion of diagnostic testing capacity on an unprecedented scale. The rapid evolution of technologies that allowed testing to move out of traditional laboratories and into point-of-care testing centers and the home transformed the diagnostic landscape. Four years later, with the end of the formal public health emergency but continued global circulation of the virus, it is important to take a fresh look at available SARS-CoV-2 testing technologies and consider how they should be used going forward. This review considers current use case scenarios for SARS-CoV-2 antigen, nucleic acid amplification, and immunologic tests, incorporating the latest evidence for analytical/clinical performance characteristics and advantages/limitations for each test type to inform current debates about how tests should or should not be used.

The use of peptides for immunodiagnosis of human Chagas disease.

Chagas disease, caused by the protozoa Trypanosoma cruzi, continues to be a serious public health problem in Latin America, worsened by the limitations in its detection. Given the importance of developing new diagnostic methods for this disease, the present review aimed to verify the number of publications dedicated to research on peptides that demonstrate their usefulness in serodiagnosis. To this end, a bibliographic survey was conducted on the PubMed platform using the keyword "peptide" or "epitope" combined with "Chagas disease" or "Trypanosoma cruzi"; "diagno*" or "serodiagnosis" or "immunodiagnosis", without period restriction. An increasing number of publications on studies employing peptides in ELISA and rapid tests assays was verified, which confirms the expansion of research in this field. It is possible to observe that many of the peptides tested so far originate from proteins widely used in the diagnosis of Chagas, and many of them are part of commercial tests developed. In this sense, as expected, promising results were obtained for several peptides when tested in ELISA, as many of them exhibited sensitivity and specificity values above 90%. Furthermore, some peptides have been tested in several studies, confirming their diagnostic potential. Despite the promising results observed, it is possible to emphasize the need for extensive testing of peptides, using different serological panels, in order to confirm their potential. The importance of producing an effective assay capable of detecting the clinical stages of the disease, as well as new immunogenic antigens that enable new serological diagnostic tools for Chagas disease, is evident.

Recombinant multiepitope proteins expressed in Escherichia coli cells and their potential for immunodiagnosis.

Recombinant multiepitope proteins (RMPs) are a promising alternative for application in diagnostic tests and, given their wide application in the most diverse diseases, this review article aims to survey the use of these antigens for diagnosis, as well as discuss the main points surrounding these antigens. RMPs usually consisting of linear, immunodominant, and phylogenetically conserved epitopes, has been applied in the experimental diagnosis of various human and animal diseases, such as leishmaniasis, brucellosis, cysticercosis, Chagas disease, hepatitis, leptospirosis, leprosy, filariasis, schistosomiasis, dengue, and COVID-19. The synthetic genes for these epitopes are joined to code a single RMP, either with spacers or fused, with different biochemical properties. The epitopes' high density within the RMPs contributes to a high degree of sensitivity and specificity. The RMPs can also sidestep the need for multiple peptide synthesis or multiple recombinant proteins, reducing costs and enhancing the standardization conditions for immunoassays. Methods such as bioinformatics and circular dichroism have been widely applied in the development of new RMPs, helping to guide their construction and better understand their structure. Several RMPs have been expressed, mainly using the Escherichia coli expression system, highlighting the importance of these cells in the biotechnological field. In fact, technological advances in this area, offering a wide range of different strains to be used, make these cells the most widely used expression platform. RMPs have been experimentally used to diagnose a broad range of illnesses in the laboratory, suggesting they could also be useful for accurate diagnoses commercially. On this point, the RMP method offers a tempting substitute for the production of promising antigens used to assemble commercial diagnostic kits.

Immunoinformatics-Based Designing of Novel and Potent Multi-Epitope PSA D15 and Cag11 Immunogens for Helicobacter pylori Immunodiagnostic Assay Development.

Helicobacter pylori (H. pylori) strain is the most genetically diverse pathogenic bacterium and now alarming serious human health concern ranging from chronic gastritis to gastric cancer and human death all over the world. Currently, the majority of commercially available diagnostic assays for H. pylori is a challenging task due to the heterogeneity of virulence factors in various geographical regions. In this concern, designing of universal multi-epitope immunogenic biomarker targeted for all H. pylori strains would be crucial to successfully immunodiagnosis assay and vaccine development for H. pylori infection. Hence, the present study aimed to explore the potential immunogenic epitopes of PSA D15 and Cag11 proteins of H. pylori, using immunoinformatics web tools in order to design novel immune-reactive multi-epitope antigens for enhanced immunodiagnosis in humans. Through an in silico immunoinformatics approach, high-ranked B-cell, MHC-I, and MHC-II epitopes of PSA D15 and Cag11 proteins were predicted, screened, and selected. Subsequently, a novel multi-epitope PSA D15 and Cag11 antigens were designed by fused the high-ranked B-cell, MHC-I, and MHC-II epitopes and 50S ribosomal protein L7/L12 adjuvant using linkers. The antigenicity, solubility, physicochemical properties, secondary and tertiary structures, 3D model refinement, and validations were carried. Furthermore, the designed multi-epitope antigens were subjected to codon adaptation and in silico cloning, immune response simulation, and molecular docking with receptor molecules. A novel, stable multi-epitope PSA D15 and Cag11 H. pylori antigens were developed and immune simulation of the designed antigens showed desirable levels of immunological response. Molecular docking of designed antigens with immune receptors (B-cell, MHC-I, MHC-II, and TLR-2/4) revealed robust interactions and stable binding affinity to the receptors. The codon optimized and in silico cloned showed that the designed antigens were successfully expressed (CAI value of 0.95 for PSA D15 and 1.0 for Cag11) after inserted into pET-32ba (+) plasmid of the E. coli K12 strain. In conclusion, this study revealed that the designed multi-epitope antigens have a huge immunological potential candidate biomarker and useful in developing immunodiagnostic assays and vaccines for H. pylori infection.

Evaluation of the "Burgenland PREvention trial of colorectal cancer Disease with ImmunologiCal Testing" (B-PREDICT)-a population-based colorectal cancer screening program.

BACKGROUND: The colorectal cancer (CRC) screening program B-PREDICT is a population based invited two stage screening project using a faecal immunochemical test (FIT) for initial screening followed by a colonoscopy for those with a positive FIT. B-PREDICT was compared with the opportunistic screening colonoscopy (OPP-COL), performed in course of the nationwide screening program. METHODS: Within B-PREDICT all residents of the Austrian federal state Burgenland, aged between 40 and 80 are annually invited to FIT testing. All individuals who underwent initial colonoscopy in Burgenland between 01/2003 and 12/2014, were included in this study. Individuals from the FIT-triggered invited screening program B-PREDICT were compared with those from the non-FIT triggered OPP-COL. RESULTS: 15 133 individuals from B-PREDICT were compared to 10 045 individuals with OPP-COL. CRC detection rates were 1.34% (CI-95%, [1.15; 1.52]) in B-PREDICT compared to 0.54% in OPP-COL (95%-CI, [0.39; 0.68] p < 0.001). The decrease in the age standardized incidence rates of CRC was more pronounced in the population screened with FIT than in the general population screened with colonoscopy. Changes in incidence rates per year were -4.4% (95%-CI, [-5.1; -3.7]) vs. -1.8% (95%-CI, [-1.9; -1.6] p < 0.001). CONCLUSIONS: B-PREDICT shows a two-fold higher detection rate of CRC as well as HRA compared to OPP-COL.

Deciphering the Interaction between Neonatal Fc Receptor and Antibodies Using a Homogeneous Bioluminescent Immunoassay.

Long half-life of therapeutic Abs and Fc fusion proteins is crucial to their efficacy and is, in part, regulated by their interaction with neonatal Fc receptor (FcRn). However, the current methods (e.g., surface plasmon resonance and biolayer interferometry) for measurement of interaction between IgG and FcRn (IgG/FcRn) require either FcRn or IgG to be immobilized on the surface, which is known to introduce experimental artifacts and have led to conflicting data. To study IgG/FcRn interactions in solution, without a need for surface immobilization, we developed a novel (to our knowledge), solution-based homogeneous binding immunoassay based on NanoBiT luminescent protein complementation technology. We optimized the assay (NanoBiT FcRn assay) for human FcRn, mouse FcRn, rat FcRn, and cynomolgus FcRn and used them to determine the binding affinities of a panel of eight Abs. Assays could successfully capture the modulation in IgG/FcRn binding based on changes in Fc fragment of the Abs. We also looked at the individual contribution of Fc and F(ab)2 on the IgG/FcRn interaction and found that Fc is the main driver for the interaction at pH 6. Our work highlights the importance of using orthogonal methods to validate affinity data generated using biosensor platforms. Moreover, the simple add-and-read format of the NanoBiT FcRn assay is amenable for high-throughput screening during early Ab discovery phase.

Evaluating Responses to Gluten Challenge: A Randomized, Double-Blind, 2-Dose Gluten Challenge Trial.

BACKGROUND & AIMS: Gluten challenge is used to diagnose celiac disease (CeD) and for clinical research. Sustained gluten exposure reliably induces histologic changes but is burdensome. We investigated the relative abilities of multiple biomarkers to assess disease activity induced by 2 gluten doses, and aimed to identify biomarkers to supplement or replace histology. METHODS: In this randomized, double-blind, 2-dose gluten-challenge trial conducted in 2 US centers (Boston, MA), 14 adults with biopsy-proven CeD were randomized to 3 g or 10 g gluten/d for 14 days. The study was powered to detect changes in villous height to crypt depth, and stopped at planned interim analysis on reaching this end point. Additional end points included gluten-specific cluster of differentiation (CD)4 T-cell analysis with HLA-DQ2-gluten tetramers and enzyme-linked immune absorbent spot, gut-homing CD8 T cells, interleukin-2, symptoms, video capsule endoscopy, intraepithelial leukocytes, and tissue multiplex immunofluorescence. RESULTS: All assessments showed changes with gluten challenge. However, time to maximal change, change magnitude, and gluten dose-response relationship varied. Villous height to crypt depth, video capsule endoscopy enteropathy score, enzyme-linked immune absorbent spot, gut-homing CD8 T cells, intraepithelial leukocyte counts, and HLA-DQ2-restricted gluten-specific CD4 T cells showed significant changes from baseline at 10 g gluten only; symptoms were significant at 3 g. Symptoms and plasma interleukin-2 levels increased significantly or near significantly at both doses. Interleukin-2 appeared to be the earliest, most sensitive marker of acute gluten exposure. CONCLUSIONS: Modern biomarkers are sensitive and responsive to gluten exposure, potentially allowing less invasive, lower-dose, shorter-duration gluten ingestion. This work provides a preliminary framework for rational design of gluten challenge for CeD research. ClinicalTrials.gov number, NCT03409796.

CXCL13 in laboratory diagnosis of Lyme neuroborreliosis-the performance of the recomBead and ReaScan CXCL13 assays in human cerebrospinal fluid samples.

The chemokine CXCL13 is used as complement to serology in the diagnostics of Lyme neuroborreliosis (LNB). We evaluated and compared the semi-quantitative, cassette-based ReaScan CXCL13 assay with the quantitative recomBead CXCL13 assay using a collection of 209 cerebrospinal fluid samples. The categorical agreement between results interpreted as negative, grey zone, and positive by the two methods was 87%. The diagnostic sensitivity was higher using the recomBead assay, whereas specificity was higher using ReaScan. Few manual steps, and a short turn-around time with no batching of samples makes the ReaScan CXCL13 assay an attractive complement to serology in the diagnostics of LNB.

COVID-19 mRNA Vaccination Generates Greater Immunoglobulin G Levels in Women Compared to Men.

We investigated whether the antibody response to coronavirus disease 2019 (COVID-19) mRNA vaccination is similar in women and men. In a community cohort without prior COVID-19, first vaccine dose produced higher immunoglobulin G (IgG) levels and percent inhibition of spike-ACE2 receptor binding, a surrogate measure of virus neutralization, in women compared to men (7.0 µg/mL, 51.6% vs 3.3 µg/mL, 36.4%). After 2 doses, IgG levels remained significantly higher for women (30.4 µg/mL) compared to men (20.6 µg/mL), while percent inhibition was similar (98.4% vs 97.7%). Sex-specific antibody response to mRNA vaccination informs future efforts to understand vaccine protection and side effects.

The clinical relevance of IgM and IgA anti-pneumococcal polysaccharide ELISA assays in patients with suspected antibody deficiency.

Unlike immunoglobulin (Ig)G pneumococcal polysaccharide (PnPS)-antibodies, PnPS IgA and IgM-antibodies are not routinely determined for the assessment of immunocompetence. It is not yet known whether an isolated inability to mount a normal IgM or IgA-PnPS response should be considered a relevant primary antibody deficiency (PAD). We studied the clinical relevance of anti-PnPS IgM and IgA-assays in patients with suspected primary immunodeficiency in a large teaching hospital in 's-Hertogenbosch, the Netherlands. Serotype-specific-PnPS IgG assays were performed; subsequently, 23-valent-PnPS IgG assays (anti-PnPS IgG assays), and later anti-PnPS IgA and IgM assays, were performed in archived material (240 patients; 304 samples). Eleven of 65 pre- and six of 10 post-immunization samples from good responders to PnPS serotype-specific IgG testing had decreased anti-PnPS IgA and/or IgM titres. Of these, three pre- and no post-immunization samples were from patients previously classified as 'no PAD'. Determination of anti-PnPS IgA and IgM in addition to anti-PnPS IgG did not reduce the need for serotype-specific PnPS IgG testing to assess immunocompetence [receiver operating characteristic (ROC) analysis of post-immunization samples: anti-PnPS IgA + IgG area under the curve (AUC) = 0.80, 95% confidence interval (CI) = 0.63-0.97; anti-PnPS IgM + IgG AUC 0.80, 95% CI = 0.62-0.98; anti-PnPS IgA + IgG + IgM AUC = 0.71, 95% CI = 0.51-0.91; anti-PnPS IgG AUC = 0.93, 95% CI = 0.85-1.00]. Our data show that patients classified as having an intact antibody response based on measurement of serotype-specific PnPS IgG can still display impaired anti-PnPS IgM and IgA responses, and that the additional measurement of anti-PnPS IgA and IgM could not reduce the need for serotype-specific IgG testing. Future studies are needed to investigate the clinical relevance of potential 'specific IgA or IgM antibody deficiency' in patients with recurrent airway infections in whom no PAD could be diagnosed according to the current definitions.

Fast diagnostic test for familial Mediterranean fever based on a kinase inhibitor.

BACKGROUND AND OBJECTIVE: Familial Mediterranean fever (FMF) is the most frequent hereditary autoinflammatory disease. Its diagnosis relies on a set of clinical criteria and a genetic confirmation on identification of biallelic pathogenic MEFV variants. MEFV encodes pyrin, an inflammasome sensor. Using a kinase inhibitor, UCN-01, we recently identified that dephosphorylation of FMF-associated pyrin mutants leads to inflammasome activation. The aim of this study was to assess whether quantifying UCN-01-mediated inflammasome activation could discriminate FMF patients from healthy donors (HD) and from patients with other inflammatory disorders (OID). METHODS: Real-time pyroptosis and IL-1ß secretion were monitored in response to UCN-01 in monocytes from FMF patients (n=67), HD (n=71) and OID patients (n=40). Sensitivity and specificity of the resulting diagnostic tests were determined by receiver operating characteristic curve analyses. RESULTS: Inflammasome monitoring in response to UCN-01 discriminates FMF patients from other individuals. Pyroptosis assessment leads to a fast FMF diagnosis while combining pyroptosis and IL-1ß dosage renders UCN-01-based assays highly sensitive and specific. UCN-01-triggered monocytes responses were influenced by MEFV gene dosage and MEFV mutations in a similar way as clinical phenotypes are. CONCLUSIONS: UCN-01-based inflammasome assays could be used to rapidly diagnose FMF, with high sensitivity and specificity.

Potential false-positive reasons for SARS-CoV-2 antibody testing and its solution.

Coronavirus disease 2019 (COVID-19) has brought a huge impact on global health and the economy. Early diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is essential for epidemic prevention and control. The detection of SARS-CoV-2 antibodies is an important criterion for diagnosing COVID-19. However, SARS-CoV-2 antibody testing also has certain false positives causing confusion in clinical diagnosis. This article summarizes the causes of false-positive detection of SARS-CoV-2 antibodies in clinical practice. The results indicate that the most common endogenous interferences include rheumatoid factor, heterophile antibodies, human anti-animal antibodies, lysozyme, complement, and cross-antigens. The exogenous interference is mainly incomplete coagulation of the specimen, contamination of the specimen, and insufficient optimization of the diagnostic kit's reaction system.

Hepatitis C virus antigen detection is an appropriate test for screening and early diagnosis of hepatitis C virus infection in at-risk populations and immunocompromised hosts.

Early diagnosis remains key for effective prevention and treatment. Unfortunately, current screening with anti-hepatitis C virus antibody (anti-HCV Ab) test may have limited utility in the diagnosis of HCV infection and reinfection. This is of special concern to at-risk population, such as immunocompromised hosts and end-stage renal failure patients on hemodialysis. HCV antigen (Ag) could be useful in identifying the ongoing infection in such clinical scenarios. Hence, we aimed to study the utility of HCV Ag testing for the diagnosis of acute and chronic hepatitis C. Of 89 samples studied, 19 were from acute hepatitis C patients who were immunocompromised or were on hemodialysis, 43 were from active chronic hepatitis C patients and 27 were from patients treated for chronic hepatitis C. All samples were tested for HCV Ag using the Abbott ARCHITECT HCV Ag assay. HCV Ag was reactive in 19/19 samples from acute hepatitis C patients and 42/43 samples from active chronic hepatitis C patients. It was nonreactive in all samples from treated patients. The test showed a sensitivity and specificity of 98.4% and 100.0%, respectively. The positive and negative predictive values were 100.0% and 96.4%, respectively. The HCV antigen test has high clinical sensitivity and specificity and is useful for the diagnosis of acute and chronic hepatitis C infection in at-risk and immunocompromised patients. Its short turnaround time and relatively low cost are advantageous for use in patients on hemodialysis and other at-risk patients who require monitoring of HCV infection and reinfection.

Analysis of the diagnostic value of serum specific antibody testing for coronavirus disease 2019.

The coronavirus disease 2019 (COVID-19) pandemic has spread to various regions worldwide. As of 27 April 2020, according to real-time statistics released by the World Health Organization, there have been 84 341 confirmed cases and 4643 deaths in China, with more than 2 979 484 confirmed cases and 206 450 deaths outside China. The detection of antibodies produced during the immune response to severe acute respiratory syndrome coronavirus 2 infections has become an important laboratory method for the diagnosis of COVID-19. However, at present, a little research on these specific antibodies has been conducted. In this study, a retrospective analysis was used to explore the dynamic changes of serum immunoglobulin M (IgM) and IgG antibody and factors affecting diagnostic efficacy, so as to provide a theoretical basis for clinical diagnosis and treatment.

Vaccination versus infection with SARS-CoV-2: Establishment of a high avidity IgG response versus incomplete avidity maturation.

Avidity is defined as the binding strength of immunoglobulin G (IgG) toward its target epitope. Avidity is directly related to affinity, as both processes are determined by the best fit of IgG to epitopes. We confirm and extend data on incomplete avidity maturation of IgG toward severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleoprotein (NP), spike protein-1 (S1), and its receptor-binding domain (RBD) in coronavirus disease 2019 (COVID-19) patients. In SARS-CoV-2-infected individuals, an initial rise in avidity maturation was ending abruptly, leading to IgG of persistently low or intermediate avidity. Incomplete avidity maturation might facilitate secondary SARS-CoV-2 infections and thus prevent the establishment of herd immunity. Incomplete avidity maturation after infection with SARS-CoV-2 (with only 11.8% of cases showing finally IgG of high avidity, that is, an avidity index > 0.6) was contrasted by regular and rapid establishment of high avidity in SARS-CoV-2 naïve individuals after two vaccination steps with the BioNTech messenger RNA (mRNA) Vaccine (78% of cases with high avidity). One vaccination step was not sufficient for induction of complete avidity maturation in vaccinated SARS-CoV-2 naïve individuals, as it induced high avidity only in 2.9% of cases within 3 weeks. However, one vaccination step was sufficient to induce high avidity in individuals with previous SARS-CoV-2 infection.

The dynamics of quantitative SARS-CoV-2 antispike IgG response to BNT162b2 vaccination.

Vaccination for SARS-CoV-2 is necessary to overcome coronavirus disease 2019 (COVID-19). However, the time-dependent vaccine-induced immune response is not well understood. This study aimed to investigate the dynamics of SARS-CoV-2 antispike immunoglobulin G (IgG) response. Medical staff participants who received two sequential doses of the BNT162b2 vaccination on days 0 and 21 were recruited prospectively from the Musashino Red Cross Hospital between March and May 2021. The quantitative antispike receptor-binding domain (RBD) IgG antibody responses were measured using the Abbott SARS-CoV-2 IgGII Quant assay (cut off ≥50 AU/ml). A total of 59 participants without past COVID-19 history were continuously tracked with serum samples. The median age was 41 (22-75) years, and 14 participants were male (23.7%). The median antispike RBD IgG and seropositivity rates were 0 (0-31.1) AU/ml, 0.3 (0-39.5) AU/ml, 529.1 (48.3-8711.4) AU/ml, 18,836.9 (742.2-57,260.4) AU/ml, and 0%, 0%, 98.3%, and 100% on days 0, 3, 14, and 28 after the first vaccination, respectively. The antispike RBD IgG levels were significantly increased after day 14 from vaccination (p < 0.001) The BNT162b2 vaccination led almost all participants to obtain serum antispike RBD IgG 14 days after the first dose.

Evaluation of a rapid antigen detection test (Panbio™ COVID-19 Ag Rapid Test Device) as a point-of-care diagnostic tool for COVID-19 in a pediatric emergency department.

We evaluated the Panbio™ COVID-19 Ag Rapid Test Device as a point-of-care diagnostic tool for COVID-19 in 357 patients at a pediatric emergency department. Thirty-four patients tested positive by reverse transcription polymerase chain reaction, of which 24 were positive by the antigen assay. The sensitivity and specificity of the assay were 70.5% and 100%, respectively.

Evaluation of a chemiluminescent enzyme immunoassay-based high-throughput SARS-CoV-2 antigen assay for the diagnosis of COVID-19: The VITROS® SARS-CoV-2 Antigen Test.

A high-throughput, fully automated antigen detection test for SARS-CoV-2 is a viable alternative to reverse-transcription polymerase chain reaction (RT-qPCR) for mass screening during outbreaks. In this study, we compared RT-qPCR for viral load and the VITROS® SARS-CoV-2 Antigen Test with reference to the results of the LUMIPULSE® SARS-CoV-2 Ag Test. Of 128 nasopharyngeal swab specimens taken from patients suspected of being infected with SARS-CoV-2, 49 were positive and 79 were negative according to RT-qPCR. Consistent dose-dependent detection with VITROS® assay was successfully achieved when using nasopharyngeal swab specimens with Ct values of 32.0 or lesser, whereas the CLEIA-based LUMIPULSE® assay was able to detect lower viral loads compared with the VITROS® assay. Our results show that the performance of the VITROS® assay was satisfactory for the diagnosis of contagious COVID-19 patients in the clinical setting. Highlights The performance of the VITROS® SARS-CoV-2 Antigen Test was sufficient for the diagnosis of contagious COVID-19. This test showed high sensitivity and specificity in the detection of SARS-CoV-2 in samples with a Ct value of 32 or less.

Evaluation of immunodiagnostic tests for human gnathostomiasis using different antigen preparations of Gnathostoma spinigerum larvae against IgE, IgM, IgG, IgG1-4 and IgG1 patterns of post-treated patients.

OBJECTIVES: The aims of the study were two-fold: (1) antigen (Ag) preparation and evaluation of three antigens of Gnathostoma spinigerum infective larvae (GsL3), crude somatic antigen (CSAg), excretory-secretory antigen (ESAg) and partially purified antigens (namely P1Ag, P2Ag and P3Ag) to differentiate IgE, IgG, IgG1-4 and IgM for human gnathostomiasis diagnosis; and (2) application of the selected ELISA for following up stored sera of patients treated with ivermectin (IVM) and albendazole (ABZ). METHODS: Different antigens were analysed by antibodies of gnathostomiasis cases, other parasite infections and healthy controls using indirect ELISA to differentiate IgE, IgG, IgG1-4 and IgM. Then, prominent antigen and immunoglobulin were used in antibody predictions of gnathostomiasis cases treated with albendazole or ivermectin. RESULTS: Sensitivity of all evaluated ELISAs: IgM-, IgG-, IgG1- and IgG4-ELISA, was 100%. IgM-ELISA with CSAg and P3Ag exhibited the highest specificity of 99%. IgG-ELISA with P2Ag resulted in the highest specificity of 92.3%. IgG1-ELISA with P2Ag and P3Ag showed excellent results with 100% specificity. Finally, P2Ag evaluated IgG1 of the followed-up cases with ABZ and IVM. Decreasing antibody IgG1 levels were mostly found in both treatments at Month 9 and long follow-up was over 12 months. A Gnathostoma worm was extracted from each two treated patients. CONCLUSIONS: Using IgG1-ELISA against P2Ag and P3Ag gave excellent results with 100% sensitivity and specificity. These tests can be an alternative to immunoblotting for gnathostomiasis. IgG1 decreased at least 9 months in most cases, so long-term treatment should be performed over 1 year.