scholarly journals Performance Comparison of a Flow Cytometry-based and Two Commercial Chemiluminescent Immunoassays for Detection and Quantification of Antibodies Binding to SARS-CoV-2 Spike Protein

2021 ◽  
Author(s):  
Arantxa Valdivia ◽  
Fabián Tarín ◽  
María Jesús Alcaraz ◽  
Paula Piñero ◽  
Ignacio Torres ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Performance Comparison ◽  
Antibody Isotype ◽  
Jurkat T Cells ◽  
Specific Antibodies ◽  
Clinical Sensitivity ◽  
Percent Agreement ◽  
Antibody Levels ◽  
Roche Diagnostics ◽  
Detection And Quantification

The performance of a laboratory-developed quantitative IgG/IgA flow cytometry-based immunoassay (FCI) using Jurkat T cells stably expressing full-length native S protein was compared against Elecsys® electrochemiluminiscent (ECLIA) Anti-SARS-CoV-2 S (Roche Diagnostics, Pleasanton, CA, USA), and LIAISON® SARS-CoV-2 TrimericS IgG chemiluminiscent assay (CLIA) (Diasorin S.p.a, Saluggia, IT) for detection and quantitation of SARS-CoV-2-specific antibodies. A total of 225 serum/plasma specimens from 120 acute or convalescent COVID-19 individuals were included. Overall, IgG/IgA-FCI yielded the highest number of positives (n=179), followed by IgA-FCI (n=177), Roche ECLIA (n=175), IgG-FCI (n=172) and Diasorin CLIA (n=154). Positive percent agreement between FCI and compared immunoassays was highest for Roche ECLIA, ranging from 96.1% (IgG/IgA-FCI) to 97.7% (IgG-FCI), whereas negative percent agreement was higher between FCI and Diasosin CLIA, regardless of antibody isotype. A strong correlation (Rho:0.6-0.8) was found between IgG-FCI or IgA-FCI levels and antibodies quantified by Roche ECLIA and Diasorin CLIA. The trajectory of antibody levels delineated by the different immunoassays in 22 of patients with sequential specimens (>=3) was frequently discordant, with the exception of IgG and IgA determined by FCI assay and to a lesser extent antibodies quantified by Roche ECLIA and Diasorin CLIA. The data suggest that FCI may outperform Roche ECLIA and Diasorin CLIA in terms of clinical sensitivity for serological diagnosis of SARS-CoV-2 infection.

scholarly journals Performance of a flow cytometry-based immunoassay for detection of antibodies binding to SARS-CoV-2 spike protein

2022 ◽  
Vol 12 (1) ◽  
Author(s):  
Arantxa Valdivia ◽  
Fabián Tarín ◽  
María Jesús Alcaraz ◽  
Paula Piñero ◽  
Ignacio Torres ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
Spike Protein ◽  
Antibody Isotype ◽  
Jurkat T Cells ◽  
Specific Antibodies ◽  
Clinical Sensitivity ◽  
Percent Agreement ◽  
Roche Diagnostics ◽  
Positive Results

AbstractThe performance of a laboratory-developed IgG/IgA flow cytometry-based immunoassay (FCI) using Jurkat T cells stably expressing full-length native S protein was compared against Elecsys electrochemiluminiscent (ECLIA) Anti-SARS-CoV-2 S (Roche Diagnostics, Pleasanton, CA, USA), and Liaison SARS-CoV-2 TrimericS IgG chemiluminiscent assay (CLIA) (Diasorin S.p.a, Saluggia, IT) for detection of SARS-CoV-2-specific antibodies. A total of 225 serum/plasma specimens from 120 acute or convalescent COVID-19 individuals were included. Overall, IgG/IgA-FCI yielded the highest number of positives (n = 179), followed by IgA-FCI (n = 177), Roche ECLIA (n = 175), IgG-FCI (n = 172) and Diasorin CLIA (n = 154). For sera collected early after the onset of symptoms (within 15 days) IgG/IgA-FCI also returned the highest number of positive results (52/72; 72.2%). Positive percent agreement between FCI and compared immunoassays was highest for Roche ECLIA, ranging from 96.1 (IgG/IgA-FCI) to 97.7% (IgG-FCI), whereas negative percent agreement was higher between FCI and Diasosin CLIA, regardless of antibody isotype. The data suggest that FCI may outperform Roche ECLIA and Diasorin CLIA in terms of clinical sensitivity for serological diagnosis of SARS-CoV-2 infection.

scholarly journals A flexible, pan-species, multi-antigen platform for the detection and monitoring of SARS-CoV-2-specific antibody responses

2021 ◽  
Author(s):  
Huifeng Shen ◽  
David Forgacs ◽  
Digantkumar Chapla ◽  
Kelley W. Moremen ◽  
Lance Wells ◽  
...  
Keyword(s):  
Host Species ◽  
Unmet Need ◽  
Reference Points ◽  
Antibody Isotype ◽  
Neutralization Activity ◽  
Specific Antibodies ◽  
E Coli ◽  
Specimen Type ◽  
Antigen Assay ◽  
Antibody Levels

AbstractThe SARS-CoV-2 pandemic and the vaccination effort that is ongoing has created an unmet need for accessible, affordable, flexible and precise platforms for monitoring the induction, specificity and maintenance of virus-specific immune responses. Herein we validate a multiplex (Luminex-based) assay capable of detecting SARS-CoV-2-specific antibodies irrespective of host species, antibody isotype, and specimen type (e.g. plasma, serum, saliva or blood spots). The well-established precision of Luminex-based assays provides the ability to follow changes in antibody levels over time to many antigens, including multiple permutations of the most common SARS-CoV-2 antigens. This platform can easily measure antibodies known to correlate with neutralization activity as well as multiple non-SARS-CoV-2 antigens such as vaccines (e.g. Tetanus toxoid) or those from frequently encountered agents (influenza), which serve as stable reference points for quantifying the changing SARS-specific responses. All of the antigens utilized in our study can be made in-house, many in E. coli using readily available plasmids. Commercially sourced antigens may also be incorporated and newly available antigen variants can be rapidly produced and integrated, making the platform adaptable to the evolving viral strains in this pandemic.Brief SummaryA multi-antigen assay for monitoring SARS-CoV-2-specific antibodies irrespective of host species, antibody isotype, and specimen type was developed.

scholarly journals 421. If at first you do not succeed…. Repeat SARS-COV2 PCR testing

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S277-S278
Author(s):  
Stephanie M Shea ◽  
Gopi Patel ◽  
Sarah Schaefer ◽  
Michael D Nowak ◽  
Emilia Mia Sordillo ◽  
...  
Keyword(s):  
Clinical Symptoms ◽  
Patient Characteristics ◽  
Nucleic Acid Amplification ◽  
Duration Of Symptoms ◽  
Radiographic Findings ◽  
Repeat Testing ◽  
Clinical Sensitivity ◽  
Viral Transport Medium ◽  
Minimum Interval ◽  
Roche Diagnostics

Abstract Background Nucleic Acid Amplification Tests (NAATs) of nasopharyngeal specimens (NPS) have become standard for diagnosis of SARS-COV2. IDSA guidelines suggest repeat testing after 24–48 h when initially negative and clinical suspicion persists. We characterized patients from whom initial NPS were NAAT-negative, but repeats were NAAT-positive, in order to identify which patients might benefit from repeat NAAT for SARS-CoV-2, and the appropriate interval. Methods We conducted an IRB-approved retrospective review of laboratory and electronic medical record data for all patients evaluated for SARS-CoV-2 infection at the Mount Sinai Health System, whose initial NAATs were done between March 16 – March 30, 2020, and who were retested within one month. NAATs were performed on NPS in viral transport medium using the Roche Diagnostics cobas® 6800 SARS-CoV-2 Test. Baseline patient characteristics, clinical and radiographic findings were identified. Results Of 235 patients eligible for inclusion, 172 (70.5%) were initially NAAT-negative, and 118 (68.6%) remained NAAT-negative over 1 month follow up. 54 (31.4%) converted to NAAT-positive over the next 1-month. Of patients who became NAAT-positive, 31 (57.4%) were inpatients who converted results within a single admission; the average interval was 6d 7h between the NAAT-negative and NAAT-positive results, and the minimum interval was 10.5 h. Symptoms examined for correlation for conversion to NAAT-positive were: fever, cough, shortness of breath, and combined nausea/vomiting/diarrhea. Duration of symptoms reported at triage did not appear to affect time to conversion to NAAT-positive. No individual symptom was more likely to be associated with conversion to NAAT- positive. However, time to conversion to NAAT-positive was shorter for patients with multiple symptoms. In general, chest radiography (CXR) findings correlated with NAAT results; interval to NAAT-positive was shorter for patients with worsening CXR findings. Conclusion Our data supports repeat testing in patients with multiple clinical symptoms suggestive of SARS CoV-2 infection and negative initial NP test results. Further studies are needed to determine the true clinical sensitivity and specificity of SARS-CoV-2 NAAT assays. Disclosures All Authors: No reported disclosures

scholarly journals Analysis of Specific Immunoglobulin G Subclass Antibodies for Serological Diagnosis of Echinococcosis by a Standard Enzyme-Linked Immunosorbent Assay

1998 ◽  
Vol 5 (5) ◽  
pp. 613-616 ◽  
Author(s):  
Felix Grimm ◽  
Friedrich E. Maly ◽  
Jian Lü ◽  
Roberto Llano
Keyword(s):  
Healthy Volunteers ◽  
Immunosorbent Assay ◽  
Immunoglobulin G ◽  
Serological Diagnosis ◽  
Enzyme Linked Immunosorbent Assay ◽  
Parasitic Infections ◽  
Igg Subclass ◽  
Specific Antibodies ◽  
Specific Igg4 ◽  
Antibody Levels

ABSTRACT The potential roles of specific antibodies of the different immunoglobulin G (IgG) subclasses in the serological diagnosis of cystic echinococcosis (CE) and alveolar echinococcosis (AE) were investigated by an enzyme-linked immunosorbent assay based on hydatid fluid as antigen. Specific antibodies of subclass 1 were found to be of major importance. In sera collected at the time of diagnosis (i.e., before any therapeutic intervention was initiated) they could be demonstrated in 14 of 15 sera from patients with CE and in all 12 sera from patients with AE. The most discriminatory and the most specific antibodies found in this study belonged to IgG subclass 4. Only one false-positive reaction was observed with 253 sera from healthy volunteers, and no cross-reactions occurred in 80 sera from patients with different parasitic infections. Specific IgG4 antibodies could be demonstrated in 61.0 to 66.7% (CE) or 47.6 to 66.7% (AE) of the cases. Antibody levels of IgG subclass 2 were elevated only moderately, and subclass 3 antibodies were detected in a few cases only. In addition, nonspecific reactions in sera of healthy volunteers or patients with other parasitic infections could partially be attributed to antibodies of subclasses 2 and 3.

scholarly journals IMMUNOREGULATION OF BLOOD SERUM ESTRADIOL AND PROGESTERONE LEVELS IN POSTMENOPAUSAL WOMEN

2019 ◽  
Vol 21 (5) ◽  
pp. 869-876
Author(s):  
A. N. Glushkov ◽  
E. G. Polenok ◽  
S. A. Mun ◽  
L. A. Gordeeva ◽  
V. A. Lutsenko ◽  
...  
Keyword(s):  
Blood Serum ◽  
Human Blood ◽  
Solid Phase ◽  
Biological Effects ◽  
Serum Levels ◽  
Human Blood Serum ◽  
Breast Cancer Patients ◽  
Specific Antibodies ◽  
Healthy Women ◽  
Antibody Levels

Specific antibodies against estradiol (Es) and progesterone (Pg) are known to modulate blood serum concentrations of these hormones and their biological effects after immunization of animals. It was suggested that specific IgA-Es and IgA-Pg could influence on Es and Pg levels in human blood serum. The purpose of this study was to identify the suggested correlations between serum Es and Pg and specific IgA-Es and IgA-Pg in postmenopausal healthy women (HW) and breast cancer patients (BCP). The serum levels of Es, Pg, IgA-Es and IgA-Pg were studied in 226 HW and 633 BCP by means of solid-phase immunoassay. The following results were obtained. The levels of Es in BCP (0.25 nmol/l) were higher than in HW (0.16; р < 0.0001). The levels of Pg were lower (0.79 vs 0.87; р < 0.0001), and individual Pg/Es ratios were lower (3.19 vs 6.64; р < 0.0001). Individual IgA-Pg/IgA-Es ratios correlated with decrease of Es (rs = -0.15; p = 0.029), with increase in Pg (rs = 0.38; р < 0.0001), and with increased Pg/Es ratio (rs = 0.29; р < 0.0001) in healthy women. Similar correlations were determined in BCP (correspondingly: rs = -0.14, р < 0.001; rs = 0.1, р = 0.009; rs = 0.15, р < 0.0001). The decrease of Es and increase of Pg and Pg/Es in BCP were less significant than in HW: the a quotients in regression у = ах+b (y = hormones levels and x = antibodies levels) in BCP were 3 to 4-fold lower than in HW. These peciliarities of interrelations between hormones and specific antibody levels were revealed only in ER+/PR+ BCP but not in ER+/PR- and ER-/PR- BCP. In conclusion, we have confirmed a suggestion about participation of specific antibodies in regulation of steroids levels in human blood serum. The immune regulation of hormonal status was weakened in BCP.

Pretransplant Donor-Specific Antibodies Detected by Single-Antigen Bead Flow Cytometry Are Associated With Inferior Kidney Transplant Outcomes

2010 ◽  
Vol 90 (10) ◽  
pp. 1079-1084 ◽  
Author(s):  
Neeraj Singh ◽  
Arjang Djamali ◽  
David Lorentzen ◽  
John D. Pirsch ◽  
Glen Leverson ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Kidney Transplant ◽  
Specific Antibodies ◽  
Transplant Outcomes ◽  
Single Antigen ◽  
Donor Specific Antibodies

scholarly journals The protective role of miR-223 in sepsis-induced mortality

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Dan Liu ◽  
Zhiding Wang ◽  
Huijuan Wang ◽  
Feifei Ren ◽  
Yanqin Li ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Cell Proliferation ◽  
T Cells ◽  
Protective Factor ◽  
Linear Regression Analysis ◽  
Multiple Linear Regression Analysis ◽  
Negative Control ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Jurkat T Cells

Abstract Lymphocyte apoptosis appears to play an important role in immunodysfunction in sepsis. We investigated the role of miR-223 in cell proliferation and apoptosis to identify potential target downstream proteins in sepsis. We recruited 143 patients with sepsis and 44 healthy controls from the Chinese PLA General Hospital. Flow cytometry was used to sort monocytes, lymphocytes, and neutrophils from fresh peripheral blood. A miR-223 mimic and inhibitor were used for transient transfection of Jurkat T cells. Quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) was used to assess expression of the miRNAs in cells. Western blot analysis was performed to measure protein expression. We evaluated the cell cycle and apoptosis by using flow cytometry (FCM) and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL). Expression of miR-223 was significantly higher in the survivor group than in the nonsurvivor group. Multiple linear regression analysis revealed that SOFA scores correlated negatively with miR-223 and monocyte counts, with β coefficients (95% CI) of − 0.048 (− 0.077, − 0.019) and − 47.707 (− 83.871, − 11.543), respectively. miR-223 expression also correlated negatively with the percentage of apoptosis in lymphocytes. The rate of apoptosis in the miR-223 mimic group was significantly lower than that of the negative control, with an adverse outcome observed in the miR-223 inhibitor group. We also found that miR-223 enhanced the proliferation of Jurkat T cells and that inhibiting miR-223 had an inhibitory effect on the G1/S transition. We conclude that miR-223 can serve as a protective factor in sepsis by reducing apoptosis and enhancing cell proliferation in lymphocytes by interacting with FOXO1. Potential downstream molecules are HSP60, HSP70, and HTRA.

scholarly journals Rapid and sensitive detection of SARS-CoV-2 antibodies by biolayer interferometry

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
John V. Dzimianski ◽  
Nicholas Lorig-Roach ◽  
Sara M. O’Rourke ◽  
David L. Alexander ◽  
Jacqueline M. Kimmey ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Optical Measurements ◽  
Enzyme Linked Immunosorbent Assay ◽  
High Complexity ◽  
Antibody Isotype ◽  
Biolayer Interferometry ◽  
Plasma Antibody ◽  
Quantitative Results ◽  
Lateral Flow Test ◽  
Antibody Levels

AbstractSerological testing to evaluate antigen-specific antibodies in plasma is generally performed by rapid lateral flow test strips that lack quantitative results or by high complexity immunoassays that are time- and labor-intensive but provide semi-quantitative results. Here, we describe a novel application of biolayer interferometry for the rapid detection of antigen-specific antibody levels in plasma samples, and demonstrate its utility for quantification of SARS-CoV-2 antibodies. Our biolayer interferometry immunosorbent assay (BLI-ISA) utilizes single-use biosensors in an automated “dip-and-read” format, providing real-time optical measurements of antigen loading, plasma antibody binding, and antibody isotype detection. Complete semi-quantitative results are obtained in less than 20 min. BLI-ISA meets or exceeds the performance of high complexity methods such as Enzyme-Linked Immunosorbent Assay (ELISA) and Chemiluminescent Immunoassay. Importantly, our method can be immediately implemented on existing BLI platforms for urgent COVID-19 studies, such as serosurveillance and the evaluation of vaccine candidates. In a broader sense, BLI-ISA can be developed as a novel diagnostic platform to evaluate antibodies and other biomolecules in clinical specimens.

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