scholarly journals A 10-Minute “Mix and Read” Antibody Assay for SARS-CoV-2

Viruses ◽  
2021 ◽  
Vol 13 (2) ◽  
pp. 143
Author(s):  
Juuso Rusanen ◽  
Lauri Kareinen ◽  
Lev Levanov ◽  
Sointu Mero ◽  
Sari H. Pakkanen ◽  
...  
Keyword(s):  
Antibody Response ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Neutralizing Antibody ◽  
Antibody Detection ◽  
Diagnostic Tools ◽  
Human Antibody ◽  
Protein L ◽  
Specific Patient ◽  
Set Up

Accurate and rapid diagnostic tools are needed for management of the ongoing coronavirus disease 2019 (COVID-19) pandemic. Antibody tests enable detection of individuals past the initial phase of infection and help examine vaccine responses. The major targets of human antibody response in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are the spike glycoprotein (SP) and nucleocapsid protein (NP). We have developed a rapid homogenous approach for antibody detection termed LFRET (protein L-based time-resolved Förster resonance energy transfer immunoassay). In LFRET, fluorophore-labeled protein L and antigen are brought to close proximity by antigen-specific patient immunoglobulins of any isotype, resulting in TR-FRET signal. We set up LFRET assays for antibodies against SP and NP and evaluated their diagnostic performance using a panel of 77 serum/plasma samples from 44 individuals with COVID-19 and 52 negative controls. Moreover, using a previously described SP and a novel NP construct, we set up enzyme linked immunosorbent assays (ELISAs) for antibodies against SARS-CoV-2 SP and NP. We then compared the LFRET assays with these ELISAs and with a SARS-CoV-2 microneutralization test (MNT). We found the LFRET assays to parallel ELISAs in sensitivity (90–95% vs. 90–100%) and specificity (100% vs. 94–100%). In identifying individuals with or without a detectable neutralizing antibody response, LFRET outperformed ELISA in specificity (91–96% vs. 82–87%), while demonstrating an equal sensitivity (98%). In conclusion, this study demonstrates the applicability of LFRET, a 10-min “mix and read” assay, to detection of SARS-CoV-2 antibodies.

scholarly journals Rapid homogeneous assay for detecting antibodies against SARS-CoV-2

2020 ◽  
Author(s):  
Juuso Rusanen ◽  
Lauri Kareinen ◽  
Lev Levanov ◽  
Sointu Mero ◽  
Sari H. Pakkanen ◽  
...  
Keyword(s):  
Antibody Response ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Neutralizing Antibody ◽  
Antibody Detection ◽  
Diagnostic Tools ◽  
Human Antibody ◽  
Protein L ◽  
Specific Patient ◽  
Set Up

AbstractAccurate and rapid diagnostic tools are needed for management of the ongoing coronavirus disease 2019 (COVID-19) pandemic. Antibody tests enable detection of individuals past the initial phase of infection and will help to examine possible vaccine responses. The major targets of human antibody response in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are the spike glycoprotein (S) and nucleocapsid protein (N). We have developed a rapid homogenous approach for antibody detection termed LFRET (protein L-based time-resolved Förster resonance energy transfer immunoassay). In LFRET, fluorophore-labeled protein L and antigen are brought to close proximity by antigen-specific patient immunoglobulins of any isotype, resulting in TR-FRET signal generation.We set up LFRET assays for antibodies against S and N and evaluated their diagnostic performance using a panel of 77 serum/plasma samples from 44 individuals with COVID-19 and 52 negative controls. Moreover, using a previously described S construct and a novel N construct, we set up enzyme linked immunosorbent assays (ELISAs) for antibodies against SARS-CoV-2 S and N. We then compared the LFRET assays with these enzyme immunoassays and with a SARS-CoV-2 microneutralization test (MNT).We found the LFRET assays to parallel ELISAs in sensitivity (90-95% vs. 90-100%) and specificity (100% vs. 94-100%). In identifying individuals with or without a detectable neutralizing antibody response, LFRET outperformed ELISA in specificity (91-96% vs. 82-87%), while demonstrating an equal sensitivity (98%).In conclusion, this study demonstrates the applicability of LFRET, a 10-minute ‘mix and read’ assay, to detection of SARS-CoV-2 antibodies.

scholarly journals Urine and Free Immunoglobulin Light Chains as Analytes for Serodiagnosis of Hantavirus Infection

Viruses ◽  
2019 ◽  
Vol 11 (9) ◽  
pp. 809 ◽  
Author(s):  
Hepojoki ◽  
Kareinen ◽  
Strandin ◽  
Vaheri ◽  
Holthöfer ◽  
...  
Keyword(s):  
Point Of Care ◽  
Resonance Energy ◽  
Kidney Injury ◽  
Hemorrhagic Fever ◽  
Disease Diagnosis ◽  
Antibody Detection ◽  
Light Chains ◽  
Hantavirus Infection ◽  
Protein L ◽  
Donor And Acceptor

Rapid point-of-care testing is a megatrend in infectious disease diagnosis. We have introduced a homogeneous immunoassay concept, which is based on the simultaneous binding of antigen and protein L to a given immunoglobulin molecule. The complex formation is detected utilizing time-resolved Förster resonance energy transfer between antigen-attached donor and acceptor-labeled protein L, hence the name LFRET. Here, we demonstrate that urine can be used as a sample matrix in LFRET-based serodiagnostics. We studied urine samples collected during the hospitalization and recovery of patients with acute Puumala orthohantavirus (PUUV) infection. We compared PUUV antibody-specific LFRET signals in urine to those in plasma, and found excellent correlation in the test outcomes The LFRET test from urine was positive in 40/40 patients with acute PUUV infection. PUUV causes a mild form of hemorrhagic fever with renal syndrome, characterized by acute kidney injury and proteinuria. Immunofluorescence and western blotting demonstrated PUUV-IgG and -IgA in urine, however, the presence of intact immunoglobulins did not fully explain the LFRET signals. We purified free light chains (FLCs) from both urine and serum of healthy volunteers and patients with acute PUUV infection, and verified the presence of antigen-specific FLCs. Antigen-specific FLCs provide a new means for non-invasive antibody detection and disease diagnosis.

scholarly journals Recent Developments of Electrochemical and Optical Biosensors for Antibody Detection

2019 ◽  
Vol 21 (1) ◽  
pp. 134 ◽  
Author(s):  
Wei Xu ◽  
Daniel Wang ◽  
Derek Li ◽  
Chung Chiun Liu
Keyword(s):  
Point Of Care ◽  
Low Cost ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Disease Diagnosis ◽  
Antibody Detection ◽  
Recent Developments ◽  
Care Systems ◽  
Point Of Care Systems ◽  
Detection Of Biomarkers

Detection of biomarkers has raised much interest recently due to the need for disease diagnosis and personalized medicine in future point-of-care systems. Among various biomarkers, antibodies are an important type of detection target due to their potential for indicating disease progression stage and the efficiency of therapeutic antibody drug treatment. In this review, electrochemical and optical detection of antibodies are discussed. Specifically, creating a non-label and reagent-free sensing platform and construction of an anti-fouling electrochemical surface for electrochemical detection are suggested. For optical transduction, a rapid and programmable platform for antibody detection using a DNA-based beacon is suggested as well as the use of bioluminescence resonance energy transfer (BRET) switch for low cost antibody detection. These sensing strategies have demonstrated their potential for resolving current challenges in antibody detection such as high selectivity, low operation cost, simple detection procedures, rapid detection, and low-fouling detection. This review provides a general update for recent developments in antibody detection strategies and potential solutions for future clinical point-of-care systems.

scholarly journals Assay Concordance between SPA and TR-FRET in High-Throughput Screening

2006 ◽  
Vol 11 (6) ◽  
pp. 606-616 ◽  
Author(s):  
Oliver Von Ahsen ◽  
Anne Schmidt ◽  
Monika Klotz ◽  
Karsten Parczyk
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Statistical Significance ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Time Resolved ◽  
Detection Technology ◽  
Significant Difference ◽  
Set Up ◽  
Detection Technologies

High-throughput screening (HTS) of large chemical libraries has become the main source of new lead compounds for drug development. Several specialized detection technologies have been developed to facilitate the cost- and time-efficient screening of millions of compounds. However, concerns have been raised, claiming that different HTS technologies may produce different hits, thus limiting trust in the reliability of HTS data. This study was aimed to investigate the reliability of the authors most frequently used assay techniques: scintillation proximity assay (SPA) and homogeneous time-resolved fluorescence resonance energy transfer (TR-FRET). To investigate the data concordance between these 2 detection technologies, the authors screened a large subset of the Schering compound library consisting of 300,000 compounds for inhibitors of a nonreceptor tyrosine kinase. They chose to set up this study in realistic HTS scale to ensure statistical significance of the results. The findings clearly demonstrate that the choice of detection technology has no significant impact on hit finding, provided that assays are biochemically equivalent. Data concordance is up to 90%. The little differences in hit findings are caused by threshold setting but not by systematic differences between the technologies. The most significant difference between the compared techniques is that in the SPA format, more false-positive primary hits were obtained.

scholarly journals Single Molecule Förster Resonance Energy Transfer Studies of the Fc Region of a Human Antibody

2013 ◽  
Vol 104 (2) ◽  
pp. 71a
Author(s):  
Cathrine A. Southern ◽  
Michael T. Kelliher ◽  
Madeline E. Gemoules ◽  
Mark S. Piraino ◽  
Ramiah D. Jacks ◽  
...  
Keyword(s):  
Energy Transfer ◽  
Single Molecule ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Forster Resonance Energy Transfer ◽  
Förster Resonance Energy Transfer ◽  
Human Antibody ◽  
Förster Resonance

scholarly journals Two-step binding kinetics of tRNAGly by the glyQS T-box riboswitch and its regulation by T-box structural elements

2018 ◽  
Author(s):  
Jiacheng Zhang ◽  
Bhaskar Chetnani ◽  
Eric D. Cormack ◽  
Wei Liu ◽  
Alfonso Mondragón ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Regulatory Rna ◽  
Structural Elements ◽  
T Box ◽  
Loop Region ◽  
Rna Elements ◽  
Set Up ◽  
Trna Binding

ABSTRACTT-box riboswitches are cis-regulatory RNA elements that regulate mRNAs encoding for aminoacyl tRNA synthetases or proteins involved in amino acid biosynthesis and transport. Rather than using small molecules as their ligands, as do most riboswitches, T-box riboswitches uniquely bind tRNA and sense their aminoacylated state. Whereas the anticodon and elbow regions of the tRNA interact with Stem I, located in the 5’ portion of the T-box, sensing of the aminoacylation state involves direct binding of the NCCA sequence at the tRNA 3’ end to the anti-terminator sequence located in the 3’ portion of the T-box. However, the kinetic trajectory that describes how each of these interactions are established temporally during tRNA binding remains unclear. Using singlemolecule fluorescence resonance energy transfer (smFRET), we demonstrate that tRNA binds to the riboswitch in a two-step process, first with anticodon recognition followed by NCCA binding, with the second step accompanied by an inward motion of the 3’ portion of the T-box riboswitch relative to Stem I. By using site-specific mutants, we further show that the T-loop region of the T-box significantly contributes to the first binding step, and that the K-turn region of the T-box influences both binding steps, but with a more dramatic effect on the second binding step. Our results set up a kinetic framework describing tRNA binding by T-box riboswitches and highlight the important roles of several T-box structural elements in regulating each binding step.SIGNIFICANCEBacteria commonly use riboswitches, cis-regulatory RNA elements, to regulate the transcription or translation of the mRNAs upon sensing signals. Unlike small molecule binding riboswitches, T-box riboswitches bind tRNA and sense their aminoacylated state. T-box modular structural elements that recognize different parts of a tRNA have been identified, however, how each of these interactions is established temporally during tRNA binding remains unclear. Our study reveals that tRNA binds to the riboswitch in a two-step mechanism, with anticodon recognition first, followed by binding to the NCCA sequence at the 3’ end of the tRNA with concomitant conformational changes in the T-box. Our results also highlight the importance of the modular structural elements of the T-box in each of the binding steps.

scholarly journals Broad and potent neutralizing human antibodies to tick-borne flaviviruses protect mice from disease

2021 ◽  
Vol 218 (5) ◽  
Author(s):  
Marianna Agudelo ◽  
Martin Palus ◽  
Jennifer R. Keeffe ◽  
Filippo Bianchini ◽  
Pavel Svoboda ◽  
...  
Keyword(s):  
Antibody Response ◽  
Neutralizing Antibodies ◽  
Natural Infection ◽  
Specific Treatment ◽  
Neutralizing Antibody ◽  
Hemorrhagic Fever ◽  
Human Antibody ◽  
Lateral Ridge ◽  
Tick Borne Encephalitis Virus ◽  
Domain Iii

Tick-borne encephalitis virus (TBEV) is an emerging human pathogen that causes potentially fatal disease with no specific treatment. Mouse monoclonal antibodies are protective against TBEV, but little is known about the human antibody response to infection. Here, we report on the human neutralizing antibody response to TBEV in a cohort of infected and vaccinated individuals. Expanded clones of memory B cells expressed closely related anti-envelope domain III (EDIII) antibodies in both groups of volunteers. However, the most potent neutralizing antibodies, with IC50s below 1 ng/ml, were found only in individuals who recovered from natural infection. These antibodies also neutralized other tick-borne flaviviruses, including Langat, louping ill, Omsk hemorrhagic fever, Kyasanur forest disease, and Powassan viruses. Structural analysis revealed a conserved epitope near the lateral ridge of EDIII adjoining the EDI–EDIII hinge region. Prophylactic or early therapeutic antibody administration was effective at low doses in mice that were lethally infected with TBEV.

scholarly journals A Comprehensive View on the Human Antibody Repertoire Against Staphylococcus aureus Antigens in the General Population

2021 ◽  
Vol 12 ◽  
Author(s):  
Tanja C. Meyer ◽  
Stephan Michalik ◽  
Silva Holtfreter ◽  
Stefan Weiss ◽  
Nele Friedrich ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
General Population ◽  
Antibody Response ◽  
Dynamic Range ◽  
Population Based ◽  
Antibody Detection ◽  
Individual Response ◽  
Human Antibody ◽  
Comprehensive Overview ◽  
Suspension Array

Our goal was to provide a comprehensive overview of the antibody response to Staphylococcus aureus antigens in the general population as a basis for defining disease-specific profiles and diagnostic signatures. We tested the specific IgG and IgA responses to 79 staphylococcal antigens in 996 individuals from the population-based Study of Health in Pomerania. Using a dilution-based multiplex suspension array, we extended the dynamic range of specific antibody detection to seven orders of magnitude, allowing the precise quantification of high and low abundant antibody specificities in the same sample. The observed IgG and IgA antibody responses were highly heterogeneous with differences between individuals as well as between bacterial antigens that spanned several orders of magnitude. Some antigens elicited significantly more IgG than IgA and vice versa. We confirmed a strong influence of colonization on the antibody response and quantified the influence of sex, smoking, age, body mass index, and serum glucose on anti-staphylococcal IgG and IgA. However, all host parameters tested explain only a small part of the extensive variability in individual response to the different antigens of S. aureus.

scholarly journals Nanotechnology-Based Approaches for the Detection of SARS-CoV-2

2020 ◽  
Vol 2 ◽  
Author(s):  
Ritika Gupta ◽  
Poonam Sagar ◽  
Nitesh Priyadarshi ◽  
Sunaina Kaul ◽  
Rajat Sandhir ◽  
...  
Keyword(s):  
Point Of Care ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Cost Effective ◽  
Diagnostic Techniques ◽  
Localized Surface Plasmon ◽  
Diagnostic Tools ◽  
Diagnostic Systems ◽  
Economic Activities ◽  
Low Sensitivity

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) as a pandemic has been validated as an extreme clinical calamity and has affected several socio-economic activities globally. Proven transmission of this virus occurs through airborne droplets from an infected person. The recent upsurge in the number of infected individuals has already exceeded the number of intensive care beds available to patients. These extraordinary circumstances have elicited the need for the development of diagnostic tools for the detection of the virus and, hence, prevent the spread of the disease. Early diagnosis and effective immediate treatment can reduce and prevent an increase in the number of cases. Conventional methods of detection such as quantitative real-time polymerase chain reaction and chest computed tomography scans have been used extensively for diagnostic purposes. However, these present several challenges, including prolonged assay requirements, labor-intensive testing, low sensitivity, and unavailability of these resources in remote locations. Such challenges urgently require fast, sensitive, and accurate diagnostic techniques for the timely detection and treatment of coronavirus disease 2019 (COVID-19) infections. Point-of-care biosensors that include paper- and chip-based diagnostic systems are rapid, cost-effective, and user friendly. In this article nanotechnology-based potential biosensors for SARS-CoV-2 diagnosis are discussed with particular emphasis on a lateral flow assay, a surface-enhanced Raman scattering-based biosensor, a localized surface plasmon resonance-based biosensor, Förster resonance energy transfer, an electrochemical biosensor, and artificial intelligence-based biosensors. Several biomolecules, such as nucleic acids, antibodies/enzymes, or aptamers, can serve as potential detection molecules on an appropriate platform, such as graphene oxide, nanoparticles, or quantum dots. An effective biosensor can be developed by using appropriate combinations of nanomaterials and technologies.

scholarly journals A Protein L -Based Immunodiagnostic Approach Utilizing Time-Resolved Förster Resonance Energy Transfer

PLoS ONE ◽  
2014 ◽  
Vol 9 (9) ◽  
pp. e106432 ◽  
Author(s):  
Satu Hepojoki ◽  
Visa Nurmi ◽  
Antti Vaheri ◽  
Klaus Hedman ◽  
Olli Vapalahti ◽  
...  
Keyword(s):  
Energy Transfer ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Forster Resonance Energy Transfer ◽  
Förster Resonance Energy Transfer ◽  
Protein L ◽  
Time Resolved ◽  
Förster Resonance
Sign in / Sign up

Export Citation Format

Share Document