scholarly journals Development and performance evaluation of a rapid in-house ELISA for retrospective serosurveillance of SARS-CoV-2

PLoS ONE ◽  
2021 ◽  
Vol 16 (2) ◽  
pp. e0246346
Author(s):  
Bijon Kumar Sil ◽  
Nowshin Jahan ◽  
Md. Ahsanul Haq ◽  
Mumtarin Jannat Oishee ◽  
Tamanna Ali ◽  
...  
Keyword(s):  
Clinical Symptoms ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Serological Tests ◽  
Predictive Values ◽  
Monitoring Tools ◽  
Healthy Donors ◽  
Kappa Value ◽  
And Performance ◽  
Selection Of

Background In the ongoing pandemic situation of COVID-19, serological tests can complement the molecular diagnostic methods, and can be one of the important tools of sero-surveillance and vaccine evaluation. Aim To develop and evaluate a rapid SARS-CoV-2 specific ELISA for detection of anti-SARS-CoV2 IgG from patients’ biological samples. Methods In order to develop this ELISA, three panels of samples (n = 184) have been used: panel 1 (n = 19) and panel 2 (n = 60) were collected from RT-PCR positive patients within 14 and after 14 days of onset of clinical symptoms, respectively; whereas panel 3 consisted of negative samples (n = 105) collected either from healthy donors or pre-pandemic dengue patients. As a capturing agent full-length SARS-CoV2 specific recombinant nucleocapsid was immobilized. Commercial SARS-CoV2 IgG kit based on chemiluminescent assay was used for the selection of samples and optimization of the assay. The threshold cut-off point, inter-assay and intra-assay variations were determined. Results The incubation/reaction time was set at a total of 30 minutes with the sensitivity of 84% (95% confidence interval, CI, 60.4%, 96.6%) and 98% (95% CI, 91.1%, 100.0%), for panel 1 and 2, respectively; with overall 94.9% sensitivity (95% CI 87.5%, 98.6%). Moreover, the clinical specificity was 97.1% (95% CI, 91.9%, 99.4%) with no cross reaction with dengue samples. The overall positive and negative predictive values are 96.2% (95% CI 89.2%, 99.2%) and 96.2% (95% CI, 90.6% 99.0%), respectively. In-house ELISA demonstrated 100% positive and negative percent agreement with Elecsys Anti-SARS-CoV-2, with Cohen’s kappa value of 1.00 (very strong agreement), while comparing 13 positive and 17 negative confirmed cases. Conclusion The assay is rapid and can be applied as one of the early and retrospective sero-monitoring tools in all over the affected areas.

scholarly journals Development And Performance Evaluation of A Rapid In-House ELISA for Retrospective Serosurveillance of SARS-CoV-2

2020 ◽  
Author(s):  
Bijon Kumar Sil ◽  
Mumtarin Jannat Oishee ◽  
Md. Ahsanul Haq ◽  
Nowshin Jahan ◽  
Tamanna Ali ◽  
...  
Keyword(s):  
Clinical Signs ◽  
Blocking Agent ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Rt Pcr ◽  
Serological Tests ◽  
Predictive Values ◽  
Monitoring Tools ◽  
Kappa Value ◽  
And Performance

Background In the ongoing pandemic situation of COVID-19, serological tests can complement the molecular diagnostic methods, and can be one of the important tools of sero-surveillance and vaccine evaluation. Aim To develop and evaluate a rapid SARS-CoV-2 specific IgG ELISA with increased sensitivity and specificity. Methods In order to develop the ELISA, three panels of samples (n=184) have been used: panel 1 (n=19) and panel 2 (n=60) were collected from RT-PCR positive patients within 14 and after 14 days respectively following the onset of clinical signs of disease whereas panel 3 consisted of negative samples (n=105) collected either from healthy donors during pre-pandemic era, pandemic era or from RT-PCR negative healthy individuals. As a capturing agent full-length SARS-CoV-2 specific recombinant nucleocapsid was immobilized and blocked using blocking agent. In total of 30 samples from the panels have been tested with Elecsys Anti-SARS-CoV-2 for selecting positive and negative controls, as well as comparator assay. The threshold cut-off point, inter-assay and intra assay variations were determined. Results The assay time was set at a total of 30 minutes with the sensitivity of 84% (95% confidence interval, CI, 60.4%, 96.6%) and 98% (95% CI, 91.1%, 100.0%), for panel 1 and 2 respectively, with overall 94.9% sensitivity (95% CI 87.5%, 98.6%). Moreover, the clinical specificity is 97.1% (95% CI, 91.9%, 99.4%) with no cross-reaction with dengue sample. The overall positive and negative predictive values are 96.2% (95% CI 89.2%, 99.2%) and 96.2% (95% CI, 90.6% 99.0%) respectively. In-house ELISA demonstrated 100% positive and negative percent agreement with ROCHE (Elecsys; Anti-SARS-CoV-2), with a Cohens kappa value of 1.00 (very strong agreement), while comparing 13 positive and 17 negative confirmed cases. Conclusion The assay is rapid and can be applied as one of the early and retrospective sero-monitoring tools in all over the affected areas.

scholarly journals Evaluation of in-house ELISA and 11 commercial ELISA kits in the serological diagnosis of COVID-19

2021 ◽  
pp. 39-52
Author(s):  
Waldemar Rastawicki ◽  
Klaudia Płaza ◽  
Adam Pietrusiński
Keyword(s):  
Viral Infections ◽  
Clinical Symptoms ◽  
High Sensitivity ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Rt Pcr ◽  
Serum Samples ◽  
Serological Tests ◽  
Igm Antibodies ◽  
Epidemiological Surveys

Introduction: ELISA-Immunoassays can complement the molecular diagnostic methods, and can be one of the important tools of sero-surveillance and vaccine evaluation. The aim of the presented study was to develop in-house ELISA and evaluate 11 commercial ELISA tests for detection of anti-SARS-CoV-2 antibodies in serum samples collected from COVID patients. Methods: In total, 237 serum samples obtained from 165 people with COVID-19 with RT-PCR confirmed SARS-CoV-2 virus infection were used for the study. The specificity of the developed in-house ELISA kit was tested using 170 serum samples obtained from patients with various bacterial and viral infections. The study used an in-house ELISA and 11 commercial ELISA kits developed by various manufacturers. Results: The presented study showed high sensitivity (81.0%) and specificity (97.2%) of the developed in-house kit in relation to the RT-PCR method. The sensitivity of the inhouse test significantly increased (98.1%) when only convalescents - persons at least 3 weeks after COVID-19 were examined. Commercial ELISA kits most frequently detected IgG antibodies (from 44.9% to 89.4%), especially in samples obtained later in the disease, and the least frequent detection of IgM antibodies (from 4.2% to 42.4%). Conclusions: All the presented ELISA kits may be used in serodiagnosis of COVID-19 however the detection of antibodies in individual tests differed quite significantly and was dependent on the period of the disease, on the class of immunoglobulins and the type of antigen used. The sensitivity of serological tests in the IgG class is clearly higher when examining samples obtained at least 2-3 weeks from the onset of clinical symptoms. Searching for IgA antibodies may be useful mainly in the early phase of the disease while IgM antibodies does not provide significant additional information. In the case of asymptomatic or mild infection, the level of antibodies is low which may be the cause problems with the correct interpretation of epidemiological surveys

scholarly journals Missed opportunities for the diagnosis of Brucella infection among slaughterhouse workers at the Kumasi Abattoir, Ghana

2017 ◽  
Vol 3 (2) ◽  
pp. 58
Author(s):  
Esimebia Adjovi Amegashie ◽  
Augustina Angelina Annan ◽  
Anthony Afum-Adjei Awuah ◽  
Richard Larbi ◽  
Nicholas Addofoh ◽  
...  
Keyword(s):  
Rose Bengal ◽  
Diagnostic Methods ◽  
Enzyme Linked Immunosorbent Assay ◽  
Public Health Importance ◽  
Serological Tests ◽  
Missed Opportunities ◽  
Predictive Values ◽  
Plate Test ◽  
Kappa Value ◽  
Sensitivity Specificity

Brucellosis is a zoonotic disease in humans with its public health importance. Laboratory diagnostic methods targeting brucellosis are not performed in hospital settings across Ghana. Very little is known about the comparative diagnostic abilities of the various tests available presently. The aim of this study therefore was to evaluate and compare diagnostic performances of Rose Bengal Plate Test (RBPT), Enzyme Linked Immunosorbent Assay (ELISA) and Polymerase Chain Reaction (PCR) employed in diagnosing Brucella infection.Two hundred and twenty Abattoir workers were randomly selected in Kumasi, Ghana. Blood samples were collected, serum extracted and tested for the presence of anti-Brucella antibodies and compared among three different techniques, using ELISA, RBPT and PCR.From the 220 participants tested for antibodies against Brucella spp., 3 (1.4%), 4 (1.8%) and 21 (9.6%) were positive for Rose Bengal Plate test, anti-Brucella ELISA IgM and anti-Brucella ELISA IgG respectively. A total of 98 (44.5%) participants tested positive by PCR. The sensitivity, specificity, positive predictive value, negative predictive values and Kappa value for Rose Bengal in comparison with PCR were 66.7%, 55.8%, 2.0%, 100% and 0.013 respectively while that for ELISA IgG in comparison with PCR were 85.7%, 71.3%, 18.4%, 98.5% and 0.212 respectively.PCR yielded the highest sensitivity and specificity among the three diagnostic methods in this study and should be considered for use at strategic reference laboratories to augment existing routine serological tests for brucella performed in laboratories in Ghana.

scholarly journals The burden of congenital Chagas disease and implementation of molecular diagnostic tools in Latin America

2018 ◽  
Vol 3 (5) ◽  
pp. e001069 ◽  
Author(s):  
Albert Picado ◽  
Israel Cruz ◽  
Maël Redard-Jacot ◽  
Alejandro G Schijman ◽  
Faustino Torrico ◽  
...  
Keyword(s):  
Latin America ◽  
Chagas Disease ◽  
Diagnostic Algorithm ◽  
Diagnostic Methods ◽  
Diagnosis And Treatment ◽  
Molecular Diagnostic ◽  
Diagnostic Tools ◽  
Molecular Tests ◽  
Epidemiological Impact ◽  
And Performance

It is estimated that between 8000 and 15 000 Trypanosoma cruzi infected babies are born every year to infected mothers in Chagas disease endemic countries. Currently, poor access to and performance of the current diagnostic algorithm, based on microscopy at birth and serology at 8–12 months after delivery, is one of the barriers to congenital Chagas disease (CCD) control. Detection of parasite DNA using molecular diagnostic tools could be an alternative or complement to current diagnostic methods, but its implementation in endemic regions remains limited. Prompt diagnosis and treatment of CCD cases would have a positive clinical and epidemiological impact. In this paper, we analysed the burden of CCD in Latin America, and the potential use of molecular tests to improve access to early diagnosis and treatment of T. cruzi infected newborns.

A point-of-care diagnostic for differentiating Ebola from endemic febrile diseases

2018 ◽  
Vol 10 (471) ◽  
pp. eaat0944 ◽  
Author(s):  
David Sebba ◽  
Alexander G. Lastovich ◽  
Melody Kuroda ◽  
Eric Fallows ◽  
Joshua Johnson ◽  
...  
Keyword(s):  
Ebola Virus ◽  
Clinical Symptoms ◽  
Point Of Care ◽  
Hemorrhagic Fever ◽  
Diagnostic Methods ◽  
Outbreak Detection ◽  
Molecular Diagnostic ◽  
Clinical Samples ◽  
Live Virus ◽  
And Control

Hemorrhagic fever outbreaks such as Ebola are difficult to detect and control because of the lack of low-cost, easily deployable diagnostics and because initial clinical symptoms mimic other endemic diseases such as malaria. Current molecular diagnostic methods such as polymerase chain reaction require trained personnel and laboratory infrastructure, hindering diagnostics at the point of need. Although rapid tests such as lateral flow can be broadly deployed, they are typically not well-suited for differentiating among multiple diseases presenting with similar symptoms. Early detection and control of Ebola outbreaks require simple, easy-to-use assays that can detect and differentiate infection with Ebola virus from other more common febrile diseases. Here, we developed and tested an immunoassay technology that uses surface-enhanced Raman scattering (SERS) tags to simultaneously detect antigens from Ebola, Lassa, and malaria within a single blood sample. Results are provided in <30 min for individual or batched samples. Using 190 clinical samples collected from the 2014 West African Ebola outbreak, along with 163 malaria positives and 233 negative controls, we demonstrated Ebola detection with 90.0% sensitivity and 97.9% specificity and malaria detection with 100.0% sensitivity and 99.6% specificity. These results, along with corresponding live virus and nonhuman primate testing of an Ebola, Lassa, and malaria 3-plex assay, indicate the potential of the SERS technology as an important tool for outbreak detection and clinical triage in low-resource settings.

scholarly journals Babesiosis as a disease of people and dogs. Molecular diagnostics: a review

2008 ◽  
Vol 53 (No. 5) ◽  
pp. 229-235 ◽  
Author(s):  
B. Skotarczak
Keyword(s):  
Clinical Symptoms ◽  
Early Stage ◽  
Correct Diagnosis ◽  
Antibody Test ◽  
Diagnostic Methods ◽  
Babesia Microti ◽  
Babesia Divergens ◽  
Polymerase Chain ◽  
Etiological Agents ◽  
Selection Of

<I>Babesia</I> is the causative agent of babesiosis, a tick-borne zoonosis which has been increasingly described throughout the world. <I>Babesia microti</I> and <I>Babesia divergens</I> are the etiological agents of human babesiosis. <I>Babesia canis</I> is the principal etiological agent of canine babesiosis. Currently, the diagnostics of babesiosis is based mainly on serological methods and the immunofluorescent antibody test (IFA) is most commonly used. However, even in the acute phase of the disease, seroconversion does not always occur. Clinical symptoms, because of their unspecificity, cannot be used to make a correct diagnosis. In this situation other diagnostic methods are needed. The use of PCR (polymerase chain reaction) is the most promising of these. An advantage of this method is that it allows identification of the parasite in the early stage of disease which enables early diagnosis, implementation of therapy and avoidance of complications. However, the standardization of this technique remains to be carried out. Selection of a genetic marker for PCR is very important for the sensitivity of this technique and it is discussed in this paper.

scholarly journals Microbiological Laboratory Diagnosis of Human Brucellosis: An Overview

Pathogens ◽  
2021 ◽  
Vol 10 (12) ◽  
pp. 1623
Author(s):  
Giovanni Di Bonaventura ◽  
Silvia Angeletti ◽  
Andrea Ianni ◽  
Tommasangelo Petitti ◽  
Giovanni Gherardi
Keyword(s):  
Clinical Symptoms ◽  
Early Stage ◽  
Point Of Care ◽  
Laboratory Diagnosis ◽  
Cross Reactivity ◽  
Diagnostic Methods ◽  
Nucleic Acid Amplification ◽  
Human Brucellosis ◽  
Microbiological Analysis ◽  
Serological Tests

Brucella spp. are Gram-negative, non-motile, non-spore-forming, slow-growing, facultative intracellular bacteria causing brucellosis. Brucellosis is an endemic of specific geographic areas and, although underreported, represents the most common zoonotic infection, with an annual global incidence of 500,000 cases among humans. Humans represent an occasional host where the infection is mainly caused by B. melitensis, which is the most virulent; B. abortus; B. suis; and B. canis. A microbiological analysis is crucial to identifying human cases because clinical symptoms of human brucellosis are variable and aspecific. The laboratory diagnosis is based on three different microbiological approaches: (i) direct diagnosis by culture, (ii) indirect diagnosis by serological tests, and (iii) direct rapid diagnosis by molecular PCR-based methods. Despite the established experience with serological tests and highly sensitive nucleic acid amplification tests (NAATs), a culture is still considered the “gold standard” in the laboratory diagnosis of brucellosis due to its clinical and epidemiological relevance. Moreover, the automated BC systems now available have increased the sensitivity of BCs and shortened the time to detection of Brucella species. The main limitations of serological tests are the lack of common interpretative criteria, the suboptimal specificity due to interspecies cross-reactivity, and the low sensitivity during the early stage of disease. Despite that, serological tests remain the main diagnostic tool, especially in endemic areas because they are inexpensive, user friendly, and have high negative predictive value. Promising serological tests based on new synthetic antigens have been recently developed together with novel point-of-care tests without the need for dedicated equipment and expertise. NAATs are rapid tests that can help diagnose brucellosis in a few hours with high sensitivity and specificity. Nevertheless, the interpretation of NAAT-positive results requires attention because it may not necessarily indicate an active infection but rather a low bacterial inoculum, DNA from dead bacteria, or a patient that has recovered. Refined NAATs should be developed, and their performances should be compared with those of commercial and home-made molecular tests before being commercialized for the diagnosis of brucellosis. Here, we review and report the most common and updated microbiological diagnostic methods currently available for the laboratory diagnosis of brucellosis.

scholarly journals Diagnostic comparison of three fully automated chemiluminescent immunoassay platforms for the detection of SARS-CoV-2 antibodies

2020 ◽  
Author(s):  
Debaprasad Parai ◽  
Girish Chandra Dash ◽  
Hari Ram Choudhary ◽  
Annalisha Peter ◽  
Usha Kiran Rout ◽  
...  
Keyword(s):  
Gold Standard ◽  
Transmission Rate ◽  
High Specificity ◽  
Molecular Diagnostic ◽  
Serological Tests ◽  
Policy Makers ◽  
Kappa Value ◽  
Chemiluminescent Immunoassay ◽  
Abbott Architect ◽  
Roche Cobas

AbstractThe whole world is battling against coronavirus disease-19 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Various strategies are taken to curb the spread of the virus and to move out from the enforced lockdown stage. Serological tests are the neediest diagnostic and surveillance tool to complement the gold standard molecular diagnostic method to track down the transmission rate of SARS-CoV-2. Automated chemiluminescent immunoassay (CLIA) based analyzers become highly demanding platforms both to clinicians and policy makers for the detection anti-SARS-CoV-2 antibodies. In this study, serum from 594 patients positive for COVID-19 and 100 samples from pre-COVID cases were tested by three automated platforms: Abbott architect i2000SR, Roche cobas e411 and Yhlo iFlash 1800 and their diagnostic accuracy were compared. All three platforms showed high specificity as claimed by manufacturer. Clinical sensitivities of the machines were calculated as 64.48% (58.67-70.3) for Abbott, 80.48% (76.62-84.34) for Roche and 76.94% (72.65-81.23) for Yhlo. The Cohen’s kappa value was determined from 0.69-0.89 when inter-rater agreements were calculated. The area under the curves (AUC) values demonstrated Roche Cobas e411 as the diagnostically most accurate platform among the three CLIA analyzers.

scholarly journals Molecular Methods for Diagnosis of Viral Encephalitis

2004 ◽  
Vol 17 (4) ◽  
pp. 903-925 ◽  
Author(s):  
Roberta L. DeBiasi ◽  
Kenneth L. Tyler
Keyword(s):  
Nucleic Acid ◽  
Viral Infections ◽  
Antiviral Drug ◽  
Developed Countries ◽  
The United States ◽  
Etiologic Agent ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Predictive Values ◽  
Cns Disease

SUMMARY Hundreds of viruses cause central nervous system (CNS) disease, including meningoencephalitis and postinfectious encephalomyelitis, in humans. The cerebrospinal fluid (CSF) is abnormal in >90% of cases; however, routine CSF studies only rarely lead to identification of a specific etiologic agent. Diagnosis of viral infections of the CNS has been revolutionized by the advent of new molecular diagnostic technologies to amplify viral nucleic acid from CSF, including PCR, nucleic acid sequence-based amplification, and branched-DNA assay. PCR is ideally suited for identifying fastidious organisms that may be difficult or impossible to culture and has been widely applied for detection of both DNA and RNA viruses in CSF. The technique can be performed rapidly and inexpensively and has become an integral component of diagnostic medical practice in the United States and other developed countries. In addition to its use for identification of etiologic agents of CNS disease in the clinical setting, PCR has also been used to quantitate viral load and monitor duration and adequacy of antiviral drug therapy. PCR has also been applied in the research setting to help discriminate active versus postinfectious immune-mediate disease, identify determinants of drug resistance, and investigate the etiology of neurologic disease of uncertain cause. This review discusses general principles of PCR and reverse transcription-PCR, including qualitative, quantitative, and multiplex techniques, with comment on issues of sensitivity, specificity, and positive and negative predictive values. The application of molecular diagnostic methods for diagnosis of specific infectious entities is reviewed in detail, including viruses for which PCR is of proven efficacy and is widely available, viruses for which PCR is less widely available or for which PCR has unproven sensitivity and specificity, and nonviral entities which can mimic viral CNS disease.

scholarly journals Determination of Viral Nucleic Acid in the Human Blood

2018 ◽  
Vol 18 (4) ◽  
pp. 208-215
Author(s):  
M. A. Abdurashitov ◽  
N. A. Netesova
Keyword(s):  
Human Blood ◽  
Viral Infections ◽  
Clinical Symptoms ◽  
Viral Disease ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Reverse Transcription Reaction ◽  
Pcr Multiplex ◽  
Human Viruses

Many acute viral infections cause similar clinical symptoms, therefore, establishing the etiology of a viral disease requires the use of whole complexes of serological or PCR tests designed to detect a particular type of pathogen. Modern methods of molecular biology allow early diagnosis of viral diseases at a time when serological diagnostic methods are not yet effective. The aim of the work was to analyze molecular diagnostic methods that allow the determination of viral nucleic acids in human blood. The article presents the classification of molecular methods for the diagnosis of viral particles in clinical specimens. Methods such asin situhybridization, reverse transcription reaction (RT-PCR), nested PCR, multiplex PCR, as well as DNA microarray technology, and the method of massive parallel sequencing are considered in detail. Particular attention is paid to NGS-technologies that were used in virology almost immediately after their appearance and allowed for detection of a number of new types of human viruses (including representatives of anelloviruses, picornaviruses, polyomaviruses, etc.). The advantages and problems associated with the application of these methods in clinical practice, as well as the prospects for their improvement are discussed.

Sign in / Sign up

Export Citation Format

Share Document