scholarly journals Rapid response flow cytometric assay for the detection of antibody responses to SARS-CoV-2

2020 ◽  
Author(s):  
Dennis Lapuente ◽  
Clara Maier ◽  
Pascal Irrgang ◽  
Julian Huebner ◽  
Sophia Antonia Peter ◽  
...  
Keyword(s):  
Antibody Responses ◽  
Quantitative Detection ◽  
Converting Enzyme ◽  
Flow Cytometric Assay ◽  
Angiotensin Converting Enzyme 2 ◽  
Specific Antibodies ◽  
Diagnostic Assays ◽  
Flow Cytometric ◽  
External Standard ◽  
Specific Igg

SARS-CoV-2 has emerged as a previously unknown zoonotic coronavirus that spread worldwide causing a serious pandemic. While reliable nucleic acid-based diagnostic assays were rapidly available, there exists only a limited number of validated serological assays. Here, we evaluated a novel flow cytometric approach based on antigen-expressing HEK 293T cells to assess spike-specific IgG and IgM antibody responses. Analyses of 201 pre-COVID-19 sera proved a high assay specificity in comparison to commercially available CLIA and ELISA systems, while also revealing the highest sensitivity in specimens from PCR-confirmed SARS-CoV-2 infected patients. Additionally, a soluble Angiotensin-Converting-Enzyme 2 (ACE-2) variant was established as external standard to quantify spike-specific antibody responses on different assay platforms. In conclusion, our newly established flow cytometric assay allows sensitive and quantitative detection of SARS-CoV-2-specific antibodies, which can be easily adopted in different laboratories and does not rely on external supply of assay kits.

scholarly journals Rapid response flow cytometric assay for the detection of antibody responses to SARS-CoV-2

2020 ◽  
Author(s):  
Dennis Lapuente ◽  
Clara Maier ◽  
Pascal Irrgang ◽  
Julian Hübner ◽  
Antonia Sophia Peter ◽  
...  
Keyword(s):  
Specific Antibody ◽  
Viral Infections ◽  
Rapid Development ◽  
Antibody Responses ◽  
Quantitative Detection ◽  
Flow Cytometric Assay ◽  
Serological Tests ◽  
Diagnostic Assays ◽  
Flow Cytometric ◽  
External Standard

Abstract SARS-CoV-2 has emerged as a previously unknown zoonotic coronavirus that spread worldwide causing a serious pandemic. While reliable nucleic acid–based diagnostic assays were rapidly available, only a limited number of validated serological assays were available in the early phase of the pandemic. Here, we evaluated a novel flow cytometric approach to assess spike-specific antibody responses.HEK 293T cells expressing SARS-CoV-2 spike protein in its natural confirmation on the surface were used to detect specific IgG and IgM antibody responses in patient sera by flow cytometry. A soluble angiotensin-converting-enzyme 2 (ACE-2) variant was developed as external standard to quantify spike-specific antibody responses on different assay platforms. Analyses of 201 pre-COVID-19 sera proved a high assay specificity in comparison to commercially available CLIA and ELISA systems, while also revealing the highest sensitivity in specimens from PCR-confirmed SARS-CoV-2-infected patients. The external standard allowed robust quantification of antibody responses among different assay platforms. In conclusion, our newly established flow cytometric assay allows sensitive and quantitative detection of SARS-CoV-2-specific antibodies, which can be easily adopted in different laboratories and does not rely on external supply of assay kits. The flow cytometric assay also provides a blueprint for rapid development of serological tests to other emerging viral infections

scholarly journals SARS-CoV-2 Antibody Responses Correlate with Resolution of RNAemia But Are Short-Lived in Patients with Mild Illness

2020 ◽  
Author(s):  
Katharina Röltgen ◽  
Oliver F Wirz ◽  
Bryan A Stevens ◽  
Abigail E Powell ◽  
Catherine A Hogan ◽  
...  
Keyword(s):  
Protective Immunity ◽  
Antibody Responses ◽  
Receptor Binding Domain ◽  
Plasma Samples ◽  
Converting Enzyme ◽  
Angiotensin Converting Enzyme 2 ◽  
Specific Antibodies ◽  
Immune Clearance ◽  
Humoral Immune ◽  
Asymptomatic Individuals

SARS-CoV-2-specific antibodies, particularly those preventing viral spike receptor binding domain (RBD) interaction with host angiotensin-converting enzyme 2 (ACE2) receptor, could offer protective immunity, and may affect clinical outcomes of COVID-19 patients. We analyzed 625 serial plasma samples from 40 hospitalized COVID-19 patients and 170 SARS-CoV-2-infected outpatients and asymptomatic individuals. Severely ill patients developed significantly higher SARS-CoV-2-specific antibody responses than outpatients and asymptomatic individuals. The development of plasma antibodies was correlated with decreases in viral RNAemia, consistent with potential humoral immune clearance of virus. Using a novel competition ELISA, we detected antibodies blocking RBD-ACE2 interactions in 68% of inpatients and 40% of outpatients tested. Cross-reactive antibodies recognizing SARS-CoV RBD were found almost exclusively in hospitalized patients. Outpatient and asymptomatic individuals' serological responses to SARS-CoV-2 decreased within 2 months, suggesting that humoral protection may be short-lived.

scholarly journals A Bead-Based Flow Cytometric Assay for MonitoringYersinia pestisExposure in Wildlife

2018 ◽  
Vol 56 (7) ◽  
Author(s):  
Jeffrey C. Chandler ◽  
Laurie A. Baeten ◽  
Doreen L. Griffin ◽  
Thomas Gidlewski ◽  
Thomas J. DeLiberto ◽  
...  
Keyword(s):  
Analytical Sensitivity ◽  
Flow Cytometric Assay ◽  
Passive Hemagglutination ◽  
Content Type ◽  
Spatially Correlated ◽  
Flow Cytometric ◽  
Standard Tool ◽  
Specific Igg ◽  
Species Specific

ABSTRACTYersinia pestisis the causative agent of plague and is considered a category A priority pathogen due to its potential for high transmissibility and the significant morbidity and mortality it causes in humans.Y. pestisis endemic to the western United States and much of the world, necessitating programs to monitor for this pathogen on the landscape. Elevated human risk of plague infection has been spatially correlated with spikes in seropositive wildlife numbers, particularly rodent-eating carnivores, which are frequently in contact with the enzootic hosts and the associated arthropod vectors ofY. pestis. In this study, we describe a semiautomated bead-based flow cytometric assay developed for plague monitoring in wildlife called the F1 Luminex plague assay (F1-LPA). Based upon Luminex/Bio-Plex technology, the F1-LPA targets serological responses to the F1 capsular antigen ofY. pestisand was optimized to analyze antibodies eluted from wildlife blood samples preserved on Nobuto filter paper strips. In comparative evaluations with passive hemagglutination, the gold standard tool for wildlife plague serodiagnosis, the F1-LPA demonstrated as much as 64× improvement in analytical sensitivity for F1-specific IgG detection and allowed for unambiguous classification of IgG status. The functionality of the F1-LPA was demonstrated for coyotes and other canids, which are the primary sentinels in wildlife plague monitoring, as well as felids and raccoons. Additionally, assay formats that do not require species-specific immunological reagents, which are not routinely available for several wildlife species used in plague monitoring, were determined to be functional in the F1-LPA.

scholarly journals The Slower Antibody Response in Myelofibrosis Patients after Two Doses of mRNA SARS-CoV-2 Vaccine Calls for a Third Dose

Biomedicines ◽  
2021 ◽  
Vol 9 (10) ◽  
pp. 1480
Author(s):  
Fabio Fiorino ◽  
Anna Sicuranza ◽  
Annalisa Ciabattini ◽  
Adele Santoni ◽  
Gabiria Pastore ◽  
...  
Keyword(s):  
Healthy Subjects ◽  
Vaccine Dose ◽  
Antibody Responses ◽  
Inhibition Activity ◽  
Specific Antibodies ◽  
Slow Kinetics ◽  
Binding Inhibition ◽  
Specific Igg ◽  
The Impact ◽  
Kinetics Of

Immunization with mRNA SARS-CoV-2 vaccines has been highly recommended and prioritized in fragile subjects, including patients with myelofibrosis (MF). Available data on the vaccine immune response developed by MF patients and the impact of ruxolitinib treatment are still too fragmented to support an informed decision on a third dose for this category of subjects. Here, we show that 76% of MF patients develop spike-specific IgG after the second mRNA SARS-CoV-2 vaccine dose, but the response has a slower kinetics compared to healthy subjects, suggesting a reduced capability of their immune system to promptly react to vaccination. A reduced ACE2/RBD binding inhibition activity of spike-specific antibodies was also observed, especially in ruxolitinib-treated patients. Our results, showing slow kinetics of antibody responses in MF patients following vaccination with mRNA SARS-CoV-2 vaccines, support the need for a third vaccine dose.

scholarly journals Persistence of SARS-CoV-2-Specific Antibodies for 13 Months after Infection

Viruses ◽  
2021 ◽  
Vol 13 (11) ◽  
pp. 2313
Author(s):  
Indrė Kučinskaitė-Kodzė ◽  
Martynas Simanavičius ◽  
Aistis Šimaitis ◽  
Aurelija Žvirblienė
Keyword(s):  
Serological Test ◽  
Antibody Responses ◽  
Serological Testing ◽  
Rt Pcr ◽  
Private Company ◽  
Seropositivity Rate ◽  
S Protein ◽  
Specific Antibodies ◽  
Specific Igg ◽  
Second Wave

Background: Dynamics of antibody responses were investigated after a SARS-CoV-2 outbreak in a private company during the first wave of the pandemic. Methods: Workers of a sewing company (Lithuania) with known SARS-CoV-2 RT-PCR result during the outbreak (April 2020) were invited to participate in the study. Virus-specific IgG and IgM were monitored 2, 6 and 13 months after the outbreak via rapid IgG/IgM serological test and SARS-CoV-2 S protein-specific IgG ELISA. Results: Six months after the outbreak, 95% (CI 86–99%) of 59 previously infected individuals had virus-specific antibodies irrespective of the severity of infection. One-third of seropositive individuals had virus-specific IgM along with IgG indicating that IgM may persist for 6 months. Serological testing 13 months after the outbreak included 47 recovered individuals that remained non-vaccinated despite a wide accessibility of COVID-19 vaccines. The seropositivity rate was 83% (CI 69–91%) excluding one case of confirmed asymptomatic reinfection in this group. Between months 6 and 13, IgG levels either declined or remained stable in 31 individual and increased in 7 individuals possibly indicating an exposure to SARS-CoV-2 during the second wave of the pandemic. Conclusions: Detectable levels of SARS-CoV-2-specific antibodies persist up to 13 months after infection for the majority of the cases.

scholarly journals The slower antibody response in myelofibrosis patients after two doses of mRNA SARS-CoV-2 vaccine calls for a third dose

2021 ◽  
Author(s):  
Fabio Fiorino ◽  
Anna Sicuranza ◽  
Annalisa Ciabattini ◽  
Adele Santoni ◽  
Gabiria Pastore ◽  
...  
Keyword(s):  
Healthy Subjects ◽  
Vaccine Dose ◽  
Binding Activity ◽  
Antibody Responses ◽  
Specific Antibodies ◽  
Risk Of Mortality ◽  
Specific Igg ◽  
Open Question ◽  
The Impact ◽  
Slow Kinetic

Immunization with mRNA SARS-CoV-2 vaccines has been highly recommended and prioritized in fragile categories with higher risk of mortality after COVID-19 disease compared to healthy people, including patients with myelofibrosis (MF). Available data on the vaccine immune re-sponse developed by MF patients, and the impact of the treatment with the inhibitor of JAK-STAT signaling ruxolitimib, are still fragmented to support an informed decision for a third dose for this category of subjects. Here, we show that 76% of MF patients develop spike-specific IgG after the second vaccine dose, but the response has a slower kinetic compared to healthy subjects, suggesting a reduced capability of their immune system to promptly react to vaccina-tion. A reduced ACE2/RBD inhibition binding activity of spike-specific antibodies was also ob-served, especially in ruxolitimib treated patients. Our results contribute to answer the open question on the induction of the antibody responses in MF patients following vaccination with COVID-19 mRNA vaccines, showing a slow kinetic that support the need for a third dose of SARS-CoV-2 mRNA vaccines.

scholarly journals Tear antibodies to SARS-CoV-2: implications for transmission

2021 ◽  
Author(s):  
Kevin John Selva ◽  
Samantha K Davis ◽  
Ebene R Haycroft ◽  
Wen Shi Lee ◽  
Ester Lopez ◽  
...  
Keyword(s):  
Ocular Surface ◽  
Post Infection ◽  
Antibody Responses ◽  
Healthy Controls ◽  
Igg Antibodies ◽  
Neutralising Antibodies ◽  
Specific Antibodies ◽  
Route Of Transmission ◽  
Specific Igg ◽  
Specific Igg Antibodies

Objectives SARS-CoV-2 can be transmitted by aerosols and the ocular surface may be an important route of transmission. Little is known about protective antibody responses to SARS-CoV-2 in tears after infection or vaccination. We analysed SARS-CoV-2 specific IgG and IgA responses in human tears after either COVID-19 infection or vaccination. Methods We recruited 16 subjects with COVID-19 infection an average of 7 months previously and 15 subjects before and 2 weeks after Comirnaty (Pfizer-BioNtech) vaccination. Plasma, saliva and basal tears were collected. Pre-pandemic plasma, saliva and basal tears from 11 individuals were included as healthy controls. Antibody responses to 5 SARS-CoV-2 antigens were measured via multiplex. Results IgG antibodies to Spike and Nucleoprotein were detected in tears, saliva and plasma from subjects with prior SARS-CoV-2 infection in comparison to uninfected controls. While RBD-specific antibodies were detected in plasma, minimal RBD-specific antibodies were detected in tears and saliva. In contrast, high levels of IgG antibodies to Spike and RBD, but not Nucleoprotein, were induced in tears, saliva and plasma of subjects receiving 2 doses of the Comirnaty vaccine. Increased levels of IgA1 and IgA2 antibodies to SARS-CoV-2 antigens were detected in plasma following infection or vaccination, but were unchanged in tears and saliva. Conclusion Both infection and vaccination induce SARS-CoV-2-specific IgG antibodies in tears. RBD-specific IgG antibodies in tears were induced by vaccination but were not present 7 months post-infection. This suggests neutralising antibodies may be low in the tears late following infection.

scholarly journals Localization of Surface Immunogenic Protein on Group B Streptococcus

2001 ◽  
Vol 69 (8) ◽  
pp. 5162-5165 ◽  
Author(s):  
Stéphane Rioux ◽  
Denis Martin ◽  
Hans-Wolfgang Ackermann ◽  
Julie Dumont ◽  
Josée Hamel ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Intact Cell ◽  
Group B Streptococcus ◽  
Immunogold Electron Microscopy ◽  
Immunogenic Protein ◽  
Flow Cytometric Assay ◽  
Specific Antibodies ◽  
Septal Region ◽  
Flow Cytometric ◽  
Group B

ABSTRACT The localization and accessibility of the group B streptococcus (GBS) surface immunogenic protein (Sip) at the surface of intact GBS cells were studied by flow cytometric assay and immunogold electron microscopy. Antibodies present in pooled sera collected from mice after immunization with purified recombinant Sip efficiently recognized native Sip at the surfaces of the different GBS strains tested, which included representatives of all nine serotypes. Examination of GBS cells by immunogold electron microscopy revealed that the Sip-specific antibodies attached preferentially to polar sites and the septal region. This result confirmed that Sip is exposed at the intact-cell surface, but it also suggests that its distribution is restricted to certain regions of the cell.

scholarly journals SARS-Like Coronavirus WIV1-CoV Does Not Replicate in Egyptian Fruit Bats (Rousettus aegyptiacus)

Viruses ◽  
2018 ◽  
Vol 10 (12) ◽  
pp. 727 ◽  
Author(s):  
Neeltje van Doremalen ◽  
Alexandra Schäfer ◽  
Vineet Menachery ◽  
Michael Letko ◽  
Trenton Bushmaker ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Angiotensin Converting Enzyme ◽  
Clinical Signs ◽  
Post Inoculation ◽  
Converting Enzyme ◽  
Fruit Bats ◽  
Post Challenge ◽  
Angiotensin Converting Enzyme 2 ◽  
Specific Antibodies ◽  
Modest Increase

Severe acute respiratory syndrome (SARS)-like WIV1-coronavirus (CoV) was first isolated from Rhinolophus sinicus bats and can use the human angiotensin converting enzyme 2 (ACE2) receptor. In the current study, we investigate the ability of WIV1-CoV to infect Rousettus aegyptiacus bats. No clinical signs were observed throughout the experiment. Furthermore, only four oropharyngeal swabs and two respiratory tissues, isolated on day 3 post inoculation, were found positive for viral RNA. Two out of twelve bats showed a modest increase in coronavirus specific antibodies post challenge. In conclusion, WIV1-CoV was unable to cause a robust infection in Rousettus aegyptiacus bats.

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