Time-resolved immunofluorescence: a sensitive and specific assay for anti-HIV antibody detection in human sera

1987 ◽  
Vol 16 (4) ◽  
pp. 303-315 ◽  
Author(s):  
A. Aceti ◽  
F. Titti ◽  
P. Verani ◽  
S. Buttò ◽  
A. Pennica ◽  
...  
Keyword(s):  
Antibody Detection ◽  
Time Resolved ◽  
Specific Assay ◽  
Human Sera ◽  
Anti Hiv

scholarly journals Engineering RegulableEscherichia coliβ-Galactosidases as Biosensors for Anti-HIV Antibody Detection in Human Sera

2001 ◽  
Vol 276 (43) ◽  
pp. 40087-40095 ◽  
Author(s):  
Neus Ferrer-Miralles ◽  
Jordi X. Feliu ◽  
Stéphane Vandevuer ◽  
Annette Müller ◽  
Joaquin Cabrera-Crespo ◽  
...  
Keyword(s):  
Antibody Detection ◽  
Human Sera ◽  
Anti Hiv

scholarly journals Inexpensive Designer Antigen for Anti-HIV Antibody Detection with High Sensitivity and Specificity

2010 ◽  
Vol 17 (3) ◽  
pp. 335-341 ◽  
Author(s):  
Sheikh M. Talha ◽  
Teppo Salminen ◽  
Deepti A. Chugh ◽  
Sathyamangalam Swaminathan ◽  
Tero Soukka ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Expression System ◽  
High Sensitivity ◽  
Antibody Detection ◽  
Human Antibody ◽  
Time Resolved ◽  
Reading Frame ◽  
Diagnostic Potential ◽  
Hiv Antibodies ◽  
Anti Hiv

ABSTRACT A novel recombinant multiepitope protein (MEP) has been designed that consists of four linear, immunodominant, and phylogenetically conserved epitopes, taken from human immunodeficiency virus (HIV)-encoded antigens that are used in many third-generation immunoassay kits. This HIV-MEP has been evaluated for its diagnostic potential in the detection of anti-HIV antibodies in human sera. A synthetic MEP gene encoding these epitopes, joined by flexible peptide linkers in a single open reading frame, was designed and overexpressed in Escherichia coli. The recombinant HIV-MEP was purified using a single affinity step, yielding >20 mg pure protein/liter culture, and used as the coating antigen in an in-house immunoassay. Bound anti-HIV antibodies were detected by highly sensitive time-resolved fluorometry, using europium(III) chelate-labeled anti-human antibody. The sensitivity and specificity of the HIV-MEP were evaluated using Boston Biomedica worldwide HIV performance, HIV seroconversion, and viral coinfection panels and were found to be comparable with those of commercially available anti-HIV enzyme immunoassay (EIA) kits. The careful choice of epitopes, high epitope density, and an E. coli-based expression system, coupled with a simple purification protocol and the use of europium(III) chelate-labeled tracer, provide the capability for the development of an inexpensive diagnostic test with high degrees of sensitivity and specificity.

scholarly journals Epidemic Preparedness—Leishmania tarentolae as an Easy-to-Handle Tool to Produce Antigens for Viral Diagnosis: Application to COVID-19

2021 ◽  
Vol 12 ◽  
Author(s):  
Ilaria Varotto-Boccazzi ◽  
Alessandro Manenti ◽  
Francesca Dapporto ◽  
Louise J. Gourlay ◽  
Beatrice Bisaglia ◽  
...  
Keyword(s):  
Rapid Development ◽  
Antibody Detection ◽  
Molecular Diagnostic ◽  
Serological Tests ◽  
Leishmania Tarentolae ◽  
Cell Factories ◽  
Effective System ◽  
Human Sera ◽  
Antiviral Antibody ◽  
Viral Diagnosis

To detect and prevent emerging epidemics, discovery platforms are urgently needed, for the rapid development of diagnostic assays. Molecular diagnostic tests for COVID-19 were developed shortly after the isolation of SARS-CoV-2. However, serological tests based on antiviral antibody detection, revealing previous exposure to the virus, required longer testing phases, due to the need to obtain correctly folded and glycosylated antigens. The delay between the identification of a new virus and the development of reliable serodiagnostic tools limits our readiness to tackle future epidemics. We suggest that the protozoan Leishmania tarentolae can be used as an easy-to-handle microfactory for the rapid production of viral antigens to face emerging epidemics. We engineered L. tarentolae to express the SARS-CoV-2 receptor-binding domain (RBD) and we recorded the ability of the purified RBD antigen to detect SARS-CoV-2 infection in human sera, with a sensitivity and reproducibility comparable to that of a reference antigen produced in human cells. This is the first application of an antigen produced in L. tarentolae for the serodiagnosis of a Coronaviridae infection. On the basis of our results, we propose L. tarentolae as an effective system for viral antigen production, even in countries that lack high-technology cell factories.

scholarly journals Antibodies against Early Proteins of Human Papillomaviruses as Diagnostic Markers for Invasive Cervical Cancer

1998 ◽  
Vol 36 (2) ◽  
pp. 475-480 ◽  
Author(s):  
Wolfgang Meschede ◽  
Klaus Zumbach ◽  
Joris Braspenning ◽  
Martin Scheffner ◽  
Luis Benitez-Bribiesca ◽  
...  
Keyword(s):  
Cervical Cancer ◽  
Invasive Cervical Cancer ◽  
Human Papillomaviruses ◽  
Antibody Detection ◽  
Diagnostic Tools ◽  
Immunological Monitoring ◽  
Hpv Dna ◽  
Native Proteins ◽  
Human Sera ◽  
E6 And E7

Cervical cancer is the most prevalent tumor in developing countries and the second most frequent cancer among females worldwide. Specific human papillomaviruses (HPVs) and, most notably, HPV types 16 and 18 are recognized as being causally associated with this malignancy. Antibodies against early HPV proteins E6 and E7 have been found more often in patients with tumors than in controls. Existing peptide enzyme-linked immunosorbent assays (ELISAs) for the detection of anti-E6 and anti-E7 antibodies in human sera have low levels of sensitivity and specificity and thus are not suitable for use as diagnostic tools. Based on highly purified recombinant native proteins, we developed four sandwich ELISAs for the detection of antibodies against HPV type 16 and 18 E6 and E7 proteins. We demonstrate their sensitivities and high degrees of specificity for cervical cancer. Among a total of 501 serum specimens from unselected patients with invasive cervical cancer, 52.9% reacted positively in at least one of the four assays. In contrast, among 244 serum specimens from control subjects without cervical cancer, only 2 reactive serum specimens (0.8%) were found. For 19 of 19 antibody-positive patients, the HPV type indicated by seroreactivity was identical to the HPV DNA type found in the tumor, which also indicates a high degree of specificity for antibody detection with respect to HPV type. In a direct comparison of 72 serum specimens from patients with cervical cancer, 56% of the specimens reacted in at least one of the four protein ELISAs, whereas 40% reacted in at least one of seven peptide ELISAs covering the four antigens. These assays could be of value for the detection of invasive cervical cancer in settings in which cytology-based early tumor screening is not available, for the clinical management of patients diagnosed with cervical cancer, and for the immunological monitoring of E6 and E7 vaccination trials.

scholarly journals DEVELOPMENT OF AN IMPROVED EPSTEIN-BARR VIRUS (EBV) NEUTRALIZING ANTIBODY ASSAY TO FACILITATE DEVELOPMENT OF A PROPHYLACTIC GP350-SUBUNIT EBV VACCINE

2020 ◽  
Vol 12 (1) ◽  
pp. e2020016
Author(s):  
Hui Liu ◽  
Lorraine Gemmell ◽  
Rui Lin ◽  
Fengrong Zuo ◽  
Henry H. Balfour ◽  
...  
Keyword(s):  
High Throughput ◽  
Neutralizing Antibody ◽  
Neutralization Assay ◽  
Epidemiologic Studies ◽  
Antibody Detection ◽  
Healthy Human ◽  
Barr Virus ◽  
Human Sera ◽  
Epstein Barr ◽  
Bioanalytical Assays

No licensed vaccine is available for prevention of EBV-associated diseases, and robust, sensitive, and high-throughput bioanalytical assays are needed to evaluate immunogenicity of gp350 subunit-based candidate EBV vaccines. Here we have developed and improved analytical tools for such a vaccine’s pre-clinical and clinical validation including a gp350-specific antibody detection assay and an EBV-GFP based neutralization assay for measuring EBV specific antibodies in human donors. The sensitivity of our previously published high-throughput EBV-GFP fluorescent focus (FFA)-based neutralization assay was further improved when guinea pig complement was supplemented using a panel of healthy human sera. Anti-gp350 antibody titers, which were evaluated using an anti-gp350 IgG ELISA assay optimized for capture and detection conditions, were moderately correlated to the FFA-based neutralization titers. Overall, these sensitive, and high-throughput bioanalytical assays are capable of characterizing the serologic response to natural EBV infection, with applications in evaluating EBV antibody status in epidemiologic studies and immunogenicity of candidate gp350-subunit EBV vaccines in clinical studies.

scholarly journals Evaluation of Rapid Neutralizing Antibody Detection Test against Rabies Virus in Human Sera

2015 ◽  
Vol 43 (2) ◽  
pp. 111-116 ◽  
Author(s):  
Kieu Anh Thi Nguyen ◽  
Thu Tuyet Nguyen ◽  
Dong Vinh Nguyen ◽  
Giang Chau Ngo ◽  
Cam Nhat Nguyen ◽  
...  
Keyword(s):  
Rabies Virus ◽  
Neutralizing Antibody ◽  
Antibody Detection ◽  
Human Sera

scholarly journals Time-Resolved FRET -Based Approach for Antibody Detection – A New Serodiagnostic Concept

PLoS ONE ◽  
2013 ◽  
Vol 8 (5) ◽  
pp. e62739 ◽  
Author(s):  
Satu Saraheimo ◽  
Jussi Hepojoki ◽  
Visa Nurmi ◽  
Anne Lahtinen ◽  
Ilkka Hemmilä ◽  
...  
Keyword(s):  
Antibody Detection ◽  
Time Resolved

scholarly journals Antibody detection by agglutination–PCR (ADAP) enables early diagnosis of HIV infection by oral fluid analysis

2018 ◽  
Vol 115 (6) ◽  
pp. 1250-1255 ◽  
Author(s):  
Cheng-ting Tsai ◽  
Peter V. Robinson ◽  
Felipe de Jesus Cortez ◽  
Maria L. B. Elma ◽  
David Seftel ◽  
...  
Keyword(s):  
Oral Fluid ◽  
Detection Threshold ◽  
Population Based ◽  
Antibody Detection ◽  
Analytical Sensitivity ◽  
Sample Collection ◽  
Clinical Sensitivity ◽  
Fluid Analysis ◽  
Hiv Antibodies ◽  
Anti Hiv

Oral fluid (OF) is a highly effective substrate for population-based HIV screening efforts, as it is noninfectious and significantly easier to collect than blood. However, anti-HIV antibodies are found at far lower concentrations in OF compared with blood, leading to poor sensitivity and a longer period of time from infection to detection threshold. Thus, despite its inherent advantages in sample collection, OF is not widely used for population screening. Here we report the development of an HIV OF assay based on Antibody Detection by Agglutination–PCR (ADAP) technology. This assay is 1,000–10,000 times more analytically sensitive than clinical enzyme-linked immunoassays (EIAs), displaying both 100% clinical sensitivity and 100% specificity for detecting HIV antibodies within OF samples. We show that the enhanced analytical sensitivity enables this assay to correctly identify HIV-infected individuals otherwise missed by current OF assays. We envision that the attributes of this improved HIV OF assay can increase testing rates of at-risk individuals while enabling diagnosis and treatment at an earlier time point.

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