scholarly journals Development of Human-Murine Chimeric Immunoglobulin G for Use in the Serological Detection of Human Flavivirus and Alphavirus Antibodies

2010 ◽  
Vol 17 (10) ◽  
pp. 1617-1623 ◽  
Author(s):  
Brett A. Thibodeaux ◽  
Amanda N. Panella ◽  
John T. Roehrig
Keyword(s):  
Positive Control ◽  
Human Infection ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Variable Regions ◽  
Saint Louis ◽  
Human Control ◽  
Human Igg1 ◽  
Serological Activity ◽  
Western Equine Encephalitis

ABSTRACT Diagnosis of human arboviral infections relies heavily on serological techniques such as the immunoglobulin M (IgM) antibody capture enzyme-linked immunosorbent assay (MAC-ELISA) and the indirect IgG ELISA. Broad application of these assays is hindered by the lack of standardized positive human control sera that react with a wide variety of flaviviruses (e.g., dengue, West Nile, yellow fever, Japanese encephalitis, Saint Louis encephalitis, and Powassan viruses), or alphaviruses (e.g., Eastern equine encephalitis, Western equine encephalitis, Venezuelan equine encephalitis, and chikungunya viruses) that can cause human disease. We have created human-murine chimeric monoclonal antibodies (cMAbs) by combining the variable regions of flavivirus (6B6C-1) or alphavirus (1A4B-6) broadly cross-reactive murine MAbs (mMAbs) with the constant region of human IgG1. These cMAbs may be used as standardized reagents capable of replacing human infection-immune-positive control sera in indirect IgG ELISA for diagnosis of all human flaviviral or alphaviral infections. The IgG cMAbs secreted from plasmid-transformed Sp2/0-Ag14 cells had serological activity identical to that of the parent mMAbs, as measured by ELISA using multiple flaviviruses or alphaviruses.

scholarly journals Development of a Human-Murine Chimeric Immunoglobulin M Antibody for Use in the Serological Detection of Human Flavivirus Antibodies

2009 ◽  
Vol 16 (5) ◽  
pp. 679-685 ◽  
Author(s):  
Brett A. Thibodeaux ◽  
John T. Roehrig
Keyword(s):  
Yellow Fever Virus ◽  
Positive Control ◽  
Encephalitis Virus ◽  
Immunoglobulin M ◽  
Chimeric Antibody ◽  
Enzyme Linked Immunosorbent Assay ◽  
Variable Regions ◽  
Igm Antibody ◽  
Antibody Capture ◽  
Serological Activity

ABSTRACT Current diagnosis of human flaviviral infections relies heavily on serological techniques such as the immunoglobulin M (IgM) antibody capture enzyme-linked immunosorbent assay (MAC-ELISA). Broad application of this assay is hindered by a lack of standardized human positive-control sera that react with the wide variety of flaviviruses that can cause human disease, e.g., dengue virus (DENV), West Nile virus (WNV), yellow fever virus (YFV), Japanese encephalitis virus (JEV), and St. Louis encephalitis virus (SLEV). We have created a human-murine chimeric antibody combining the variable regions of the broadly flavivirus cross-reactive murine monoclonal antibody (MAb) 6B6C-1 and the constant region of human IgM to produce a standardized reagent capable of replacing human positive-control sera in a MAC-ELISA for the diagnosis of all human flaviviral infections. The human-murine chimeric IgM antibody secreted from plasmid-transformed Sp2/0-Ag14 cells had a level of serological activity identical to that of 6B6C-1 as measured by ELISA, immunoblotting, and MAC-ELISA for multiple members of the flavivirus genus, including WNV, SLEV, YFV, DENV, and JEV.

scholarly journals Fractionation of Membrane Components from Tachyzoite Forms ofToxoplasma gondii: Differential Recognition by Immunoglobulin M (IgM) and IgG Present in Sera from Patients with Acute or Chronic Toxoplasmosis

2000 ◽  
Vol 38 (4) ◽  
pp. 1453-1460 ◽  
Author(s):  
Mónica Giraldo ◽  
Hélia Cannizzaro ◽  
Michael A. J. Ferguson ◽  
Igor C. Almeida ◽  
Ricardo T. Gazzinelli
Keyword(s):  
Solvent Extraction ◽  
Toxoplasma Gondii ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Secondary Antibody ◽  
Igg Antibodies ◽  
Organic Solvent Extraction ◽  
Differential Recognition ◽  
Human Igg1 ◽  
Membrane Components

Tachyzoite forms of Toxoplasma gondii were subjected to a sequential organic solvent extraction, which allows fractionation of membrane components according to their degrees of hydrophobicity, yielding three fractions named F1 (most hydrophobic) to F3 (least hydrophobic). Fractions F2 (80.85% specificity and 86.95% sensitivity) and F3 (89.36% specificity and 93.61% sensitivity) gave the best results, being preferentially recognized by immunoglobulin M (IgM) and IgG in sera from patients with acute and chronic toxoplasmosis, respectively. Improved scores of specificity (100%) and sensitivity (100%) were achieved when a secondary antibody against human IgG1 instead of total IgG was employed to measure the reactivity of IgG antibodies with the F3 fraction. To purify tachyzoite antigens recognized by human IgM or IgG antibodies, the F2 or F3 fraction was loaded onto an octyl-Sepharose column and eluted with a propan-1-ol gradient. The main antigen(s) recognized by IgM or IgG eluted in a single peak from the octyl-Sepharose resin loaded with either F2 (30 to 50% propan-1-ol) or F3 (15 to 35% propan-1-ol), respectively. These semipurified fractions gave improved scores when used to detect T. gondii-specific IgM (95.7% specificity and 81.8% sensitivity) or IgG (100% specificity and 93.75% sensitivity) in an enzyme-linked immunosorbent assay. Further biochemical and immunological analyses of antigens partially purified from F2 and F3 indicate that glycoinositolphospholipids are preferentially recognized by IgM, whereas proteins of approximately 30 to 40 kDa are recognized by IgG, elicited duringT. gondii infection in humans.

scholarly journals Seroprevalence of arboviruses in Ecuador: Implications for improved surveillance

Biomédica ◽  
2021 ◽  
Vol 41 (2) ◽  
pp. 247-259
Author(s):  
Ernesto Gutiérrez-Vera ◽  
Leandro Patiño ◽  
Martha Castillo-Segovia ◽  
Víctor Mora-Valencia ◽  
Julio Montesdeoca-Agurto ◽  
...  
Keyword(s):  
Amazon Basin ◽  
Yellow Fever Virus ◽  
Encephalitis Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Igg Antibodies ◽  
Saint Louis ◽  
Equine Encephalitis Virus ◽  
Population Immunity ◽  
Vaccination History ◽  
Western Equine Encephalitis

Introduction: Arthropod-borne viruses (arboviruses) cause morbidity and mortality in humans and domestic animals worldwide. The percentage of population immunity or susceptibility to these viruses in Ecuador is unknown.Objectives: To investigate the proportion of Ecuadorian populations with IgG antibodies (Abs) (past exposure/immunity) and IgM Abs (current exposure) against flaviviruses and alphaviruses and to study the activity of these viruses in Ecuador.Materials and methods: During 2009-2011, we conducted a serosurvey for selected arboviruses in humans (n=1,842), equines (n=149), and sentinel hamsters (n=84) at two coastal locations and one in the Amazon basin (Eastern Ecuador) using enzyme-linked immunosorbent assay and hemagglutination inhibition test.Results: From 20.63% to 63.61% of humans showed IgG-antibodies for the flaviviruses: Dengue virus (DENV), yellow fever virus (YFV) Saint Louis encephalitis virus, and West Nile virus (WNV); from 4.67% to 8.63% showed IgG-Abs for the alphaviruses: Venezuelan equine encephalitis virus, eastern equine encephalitis virus, and western equine encephalitis virus. IgM-Abs were found for DENV and WNV. Equines and hamsters showed antibodies to alphaviruses in all locations; two hamsters seroconverted to YFV in the Amazonia.Conclusions: The results show a YFV vaccination history and suggest the activity of arboviruses not included in the current surveillance scheme. Enhanced arbovirus and mosquito surveillance, as well as continued YFV vaccination and evaluation of its coverage/effectiveness, are recommended.

TBE in Sweden

2020 ◽  
Keyword(s):  
Genetic Characterization ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Second Phase ◽  
Serum Samples ◽  
Tick Borne Encephalitis ◽  
Specific Immunoglobulin ◽  
Tick Borne Encephalitis Virus ◽  
First Time

Tick-borne encephalitis virus (TBEV) was isolated for the first time in Sweden in 1958 (from ticks and from 1 tick-borne encephalitis [TBE] patient).1 In 2003, Haglund and colleagues reported the isolation and antigenic and genetic characterization of 14 TBEV strains from Swedish patients (samples collected 1991–1994).2 The first serum sample, from which TBEV was isolated, was obtained 2–10 days after onset of disease and found to be negative for anti-TBEV immunoglobulin M (IgM) by enzyme-linked immunosorbent assay (ELISA), whereas TBEV-specific IgM (and TBEV-specific immunoglobulin G/cerebrospinal fluid [IgG/CSF] activity) was demonstrated in later serum samples taken during the second phase of the disease.

scholarly journals A Novel Mouse Monoclonal Antibody C42 against C-Terminal Peptide of Alpha-1-Antitrypsin

2021 ◽  
Vol 22 (4) ◽  
pp. 2141
Author(s):  
Srinu Tumpara ◽  
Elena Korenbaum ◽  
Mark Kühnel ◽  
Danny Jonigk ◽  
Beata Olejnicka ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Flow Cytometry ◽  
Western Blotting ◽  
Complex Mixture ◽  
Immunoglobulin M ◽  
Confocal Laser ◽  
Enzyme Linked Immunosorbent Assay ◽  
Dot Blot

The C-terminal-fragments of alpha1-antitrypsin (AAT) have been identified and their diverse biological roles have been reported in vitro and in vivo. These findings prompted us to develop a monoclonal antibody that specifically recognizes C-36 peptide (corresponding to residues 359–394) resulting from the protease-associated cleavage of AAT. The C-36-targeting mouse monoclonal Immunoglobulin M (IgM) antibody (containing κ light chains, clone C42) was generated and enzyme-linked immunosorbent assay (ELISA)-tested by Davids Biotechnologie GmbH, Germany. Here, we addressed the effectiveness of the novel C42 antibody in different immunoassay formats, such as dot- and Western blotting, confocal laser microscopy, and flow cytometry. According to the dot-blot results, our novel C42 antibody detects the C-36 peptide at a range of 0.1–0.05 µg and shows no cross-reactivity with native, polymerized, or oxidized forms of full-length AAT, the AAT-elastase complex mixture, as well as with shorter C-terminal fragments of AAT. However, the C42 antibody does not detect denatured peptide in SDS-PAGE/Western blotting assays. On the other hand, our C42 antibody, unconjugated as well as conjugated to DyLight488 fluorophore, when applied for immunofluorescence microscopy and flow cytometry assays, specifically detected the C-36 peptide in human blood cells. Altogether, we demonstrate that our novel C42 antibody successfully recognizes the C-36 peptide of AAT in a number of immunoassays and has potential to become an important tool in AAT-related studies.

scholarly journals Is it time to administer acellular pertussis vaccine to childbearing age/pregnant women in all areas using whole-cell pertussis vaccination schedule?

2021 ◽  
Vol 9 ◽  
pp. 251513552110158
Author(s):  
Abdoulreza Esteghamati ◽  
Shirin Sayyahfar ◽  
Yousef Alimohamadi ◽  
Sarvenaz Salahi ◽  
Mahmood Faramarzi
Keyword(s):  
Pregnant Women ◽  
Age Groups ◽  
Vaccination Schedule ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Childbearing Age ◽  
Cross Sectional ◽  
Whole Cell ◽  
Childbearing Age Women ◽  
The Mean

Background: Whole-cell pertussis (wP) vaccine administration is still advocated for children under 7 years of age in Iran. However, there is no recommendation for the administration of a dose of tetanus, diphtheria, and acellular pertussis (Tdap) vaccine to childbearing age/pregnant women in the Iranian vaccination program and it has increased the risk of infection through waning immunity during women’s childbearing age life. The study aimed to assess the levels of anti- Bordetella pertussis antibodies in childbearing age women of different ages in Iran. Methods: A cross-sectional study was conducted on a total number of 360 childbearing age women divided into six age groups, with 5-year intervals from 15 to 45 years old, in 2018–2019. Then, the levels of immunoglobulin A (IgA), immunoglobulin M (IgM), and immunoglobulin G (IgG) antibodies against B. pertussis were evaluated using enzyme-linked immunosorbent assay (ELISA). The IBM SPSS Statistics software (version 16.0) (SPSS Inc., Chicago, IL, USA) was used for data analysis. Results: The mean age of the participants was 30.01 ± 8.35 years (range 14–45 years). All the cases were IgM negative, but two IgA-positive individuals (in the age groups of 14–19 and 30–34 years) were reported. Overall, 239 (66.4%) cases were IgG positive. The mean age of IgG-positive cases was 30.37 ± 8.37 years. The IgG-positive cases were mostly in the age groups of 30–34 and 35–39 years [43 (71.1%)]. The odds of IgG positivity were 1.97. The highest odds of IgG positivity were seen in 30–34 and 35–39 years groups (2.52) and the lowest odds were seen in the 20–24 and 25–29 years groups (1.60). Using the Jonckheere–Terpstra test, the increasing trend of IgG changes in different age groups was not statistically significant (Tπ=5.78, p = 0.09). Conclusion: The infants of women of childbearing age might be prone to pertussis in countries using the wP vaccination schedule. It is suggested to administer a dose of Tdap to women before or during pregnancy to increase the immunity of their infants against this disease during early infancy.

scholarly journals Early Confirmation of Mycoplasma pneumoniae Infection by Two Short-Term Serologic IgM Examination

Diagnostics ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 353
Author(s):  
Ha Eun Jeon ◽  
Hyun Mi Kang ◽  
Eun Ae Yang ◽  
Hye Young Han ◽  
Seung Beom Han ◽  
...  
Keyword(s):  
Mycoplasma Pneumoniae ◽  
Early Stage ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Short Term ◽  
Serologic Tests ◽  
Polymerase Chain ◽  
Positive Correlations ◽  
Patient Selection Bias ◽  
Mycoplasma Pneumoniae Infection

The aim of the present study is to re-evaluate the clinical application of two-times serologic immunoglobulin M (IgM) tests using microparticle agglutination assay (MAA), an enzyme-linked immunosorbent assay (ELISA), and polymerase chain reaction (PCR) assay in diagnosing Mycoplasma pneumoniae (MP) infection. A retrospective analysis of 62 children with MP pneumonia during a recent epidemic (2019–2020) was conducted. The MAA and ELISA immunoglobulin M (IgM) and IgG measurements were conducted twice at admission and around discharge, and MP PCR once at presentation. Diagnostic rates in each test were calculated at presentation and at discharge. The seroconverters were 39% (24/62) of patients tested by MAA and 29% (18/62) by ELISA. At presentation, the diagnostic positive rates of MAA, ELISA, and PCR tests were 61%, 71%, and 52%, respectively. After the second examination, the rates were 100% in both serologic tests. There were positive correlations between the titers of MAA and the IgM values of ELISA. The single serologic IgM or PCR tests had limitations to select patients infected with MP in the early stage. The short-term, paired IgM serologic tests during hospitalization can reduce patient-selection bias in MP infection studies.

scholarly journals Limited Diagnostic Capacities of Two Commercial Assays for the Detection of Leptospira Immunoglobulin M Antibodies in Laos

2006 ◽  
Vol 13 (10) ◽  
pp. 1166-1169 ◽  
Author(s):  
Stuart D. Blacksell ◽  
Lee Smythe ◽  
Rattanaphone Phetsouvanh ◽  
Michael Dohnt ◽  
Rudy Hartskeerl ◽  
...  
Keyword(s):  
Diagnostic Accuracy ◽  
Immunosorbent Assay ◽  
Gold Standard ◽  
Agglutination Test ◽  
Immunoglobulin M ◽  
Microscopic Agglutination Test ◽  
Enzyme Linked Immunosorbent Assay ◽  
Diagnostic Utility

ABSTRACT The diagnostic utility of immunochromatographic (Leptotek) and enzyme-linked immunosorbent assay (ELISA; Panbio) tests for the detection of Leptospira immunoglobulin M antibodies was assessed in febrile adults admitted in Vientiane, Laos. Both tests demonstrated poor diagnostic accuracy using admission serum (Leptotek sensitivity of 47.3% and specificity of 75.5%: ELISA sensitivity of 60.9% and specificity of 65.6%) compared to the Leptospira “gold standard” microscopic agglutination test.

scholarly journals Diagnosis of Dengue Virus Infection by Detection of Specific Immunoglobulin M (IgM) and IgA Antibodies in Serum and Saliva

2003 ◽  
Vol 10 (2) ◽  
pp. 317-322 ◽  
Author(s):  
Angel Balmaseda ◽  
María G. Guzmán ◽  
Samantha Hammond ◽  
Guillermo Robleto ◽  
Carolina Flores ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Immunoglobulin M ◽  
Antibody Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Control Group ◽  
Serum Samples ◽  
Diagnostic Modality ◽  
Iga Antibodies ◽  
Specific Immunoglobulin ◽  
Feasible Alternative

ABSTRACT To evaluate alternative approaches to the serological diagnosis of dengue virus (DEN) infection, the detection of DEN-specific immunoglobulin M (IgM) and IgA antibodies in serum and saliva specimens was assessed in 147 patients with symptoms of DEN infection seen at the Ministry of Health in Nicaragua. Seventy-two serum samples were determined to be positive for anti-DEN antibodies by IgM capture enzyme-linked immunosorbent assay, the routine diagnostic procedure. Serum and saliva specimens were obtained from 50 healthy adults as additional controls. IgM was detected in the saliva of 65 of the 72 serum IgM-positive cases, 6 of the 75 serum IgM-negative cases, and none of the control group, resulting in a sensitivity of 90.3% and a specificity of 92.0% and demonstrating that salivary IgM is a useful diagnostic marker for DEN infection. Detection of IgA in serum may be another feasible alternative for the diagnosis of DEN infection, with serum IgA found in 68 (94.4%) of the IgM-positive cases. In contrast, detection of IgA in saliva was not found to be a useful tool for DEN diagnosis in the present study. Further studies of the kinetics of antibody detection in another set of 151 paired acute- and convalescent-phase serum samples showed that DEN-specific IgA antibodies were detected in more acute-phase samples than were IgM antibodies. Thus, we conclude that DEN-specific IgA in serum is a potential diagnostic target. Furthermore, given that saliva is a readily obtainable, noninvasive specimen, detection of DEN-specific salivary IgM should be considered a useful, cheaper diagnostic modality with similar sensitivity and specificity to IgM detection in serum.

scholarly journals Unreliable Measurement of Basophil Maximum Leukotriene Release with the Bühlmann CAST 2000 Enzyme-Linked Immunosorbent Assay Kit

2006 ◽  
Vol 13 (3) ◽  
pp. 420-422 ◽  
Author(s):  
S. E. Burastero ◽  
C. Paolucci ◽  
D. Breda ◽  
G. Monasterolo ◽  
R. E. Rossi ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Immunosorbent Assay ◽  
False Positive ◽  
Positive Control ◽  
Enzyme Linked Immunosorbent Assay ◽  
Positive Results

ABSTRACT The Bühlmann CAST 2000 enzyme-linked immunosorbent assay is a potentially useful assay for measuring sulfidoleukotrienes released in vitro by allergen-challenged basophils. However, we observed that the positive-control reagent yielded positive signals in cell-free systems. These false-positive results depended on using a mouse anti-FcεRI monoclonal antibody and were prevented by degranulation-inducing reagents other than mouse monoclonal antibodies.

Sign in / Sign up

Export Citation Format

Share Document