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 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 Comparison of Two Rapid Diagnostic Assays for Detection of Immunoglobulin M Antibodies to Dengue Virus

2000 ◽  
Vol 7 (1) ◽  
pp. 106-110 ◽  
Author(s):  
Shuenn-Jue L. Wu ◽  
Helene Paxton ◽  
Barbara Hanson ◽  
Cheryl G. Kung ◽  
Timothy B. Chen ◽  
...  
Keyword(s):  
Dengue Virus ◽  
False Positive ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Specific Detection ◽  
Diagnostic Assays ◽  
Igm Antibody ◽  
Elisa Format ◽  
Assay Time ◽  
Antibody Capture

ABSTRACT Two easy-to-use commercial diagnostic assays, a dipstick enzyme-linked immunosorbent assay (ELISA) (Integrated Diagnostics, Baltimore, Md.) and an immunochromatographic card assay (PanBio, Brisbane, Australia) were evaluated for detection of immunoglobulin M (IgM) antibody to dengue virus with an in-house IgM antibody capture microplate ELISA as a reference assay. The dipstick ELISA was based on the indirect-ELISA format using dengue 2 virus as the only antigen and enzyme-labeled goat anti-human IgM antibody as the detector. The total assay time was 75 min. The immunochromatographic card assay was based on the antibody capture format and separately measured both anti-dengue virus IgM and IgG in the same test. Colloidal-gold-labeled anti-dengue virus monoclonal antibody bound with dengue virus 1 to 4 antigen cocktail was the detector, and anti-human IgM and IgG were the capture antibodies. The total assay time was <10 min. Sera from 164 individuals classified as either anti-dengue virus IgM positive (94) or anti-dengue virus IgM negative (70) in the reference microplate ELISA with a dengue virus 1 to 4 antigen cocktail were tested in the two commercial assays. The dipstick ELISA missed 7 of 94 positive samples, for a sensitivity of 92.6%, while the immunochromatographic card assay missed two positive samples, for a sensitivity of 97.9%. Of the 70 negative samples, four were false positive by the dipstick ELISA and two were false positive in the immunochromatographic card assay, resulting in specificities of 94.3 and 97.1%, respectively. Both commercial assays provide sensitive and specific detection of anti-dengue virus IgM antibody and could prove useful in settings where the microplate ELISA is impractical.

scholarly journals Establishment of an Algorithm Using prM/E- and NS1-Specific IgM Antibody-Capture Enzyme-Linked Immunosorbent Assays in Diagnosis of Japanese Encephalitis Virus and West Nile Virus Infections in Humans

2015 ◽  
Vol 54 (2) ◽  
pp. 412-422 ◽  
Author(s):  
Jedhan U. Galula ◽  
Gwong-Jen J. Chang ◽  
Shih-Te Chuang ◽  
Day-Yu Chao
Keyword(s):  
West Nile Virus ◽  
Japanese Encephalitis Virus ◽  
Japanese Encephalitis ◽  
Cross Reactivity ◽  
Encephalitis Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Test Accuracy ◽  
West Nile ◽  
Igm Antibody ◽  
Antibody Capture

The front-line assay for the presumptive serodiagnosis of acute Japanese encephalitis virus (JEV) and West Nile virus (WNV) infections is the premembrane/envelope (prM/E)-specific IgM antibody-capture enzyme-linked immunosorbent assay (MAC-ELISA). Due to antibody cross-reactivity, MAC-ELISA-positive samples may be confirmed with a time-consuming plaque reduction neutralization test (PRNT). In the present study, we applied a previously developed anti-nonstructural protein 1 (NS1)-specific MAC-ELISA (NS1-MAC-ELISA) on archived acute-phase serum specimens from patients with confirmed JEV and WNV infections and compared the results with prM/E containing virus-like particle-specific MAC-ELISA (VLP-MAC-ELISA). Paired-receiver operating characteristic (ROC) curve analyses revealed no statistical differences in the overall assay performances of the VLP- and NS1-MAC-ELISAs. The two methods had high sensitivities of 100% but slightly lower specificities that ranged between 80% and 100%. When the NS1-MAC-ELISA was used to confirm positive results in the VLP-MAC-ELISA, the specificity of serodiagnosis, especially for JEV infection, was increased to 90% when applied in areas where JEV cocirculates with WNV, or to 100% when applied in areas that were endemic for JEV. The results also showed that using multiple antigens could resolve the cross-reactivity in the assays. Significantly higher positive-to-negative (P/N) values were consistently obtained with the homologous antigens than those with the heterologous antigens. JEV or WNV was reliably identified as the currently infecting flavivirus by a higher ratio of JEV-to-WNV P/N values or vice versa. In summary of the above-described results, the diagnostic algorithm combining the use of multiantigen VLP- and NS1-MAC-ELISAs was developed and can be practically applied to obtain a more specific and reliable result for the serodiagnosis of JEV and WNV infections without the need for PRNT. The developed algorithm should provide great utility in diagnostic and surveillance activities in which test accuracy is of utmost importance for effective disease intervention.

scholarly journals Technical viability of the YF MAC-HD ELISA kit for use in yellow fever-endemic regions

2021 ◽  
Vol 15 (6) ◽  
pp. e0009417
Author(s):  
Christin H. Goodman ◽  
Maurice Demanou ◽  
Mick Mulders ◽  
Jairo Mendez-Rico ◽  
Alison Jane Basile
Keyword(s):  
South America ◽  
Yellow Fever ◽  
Accurate Diagnosis ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Elisa Kit ◽  
Wash Buffer ◽  
Antibody Capture ◽  
Vaccination Campaigns ◽  
Arboviral Disease

Yellow fever (YF), an arboviral disease, affects an estimated 200,000 people and causes 30,000 deaths per year and recently has caused major epidemics in Africa and South America. Timely and accurate diagnosis of YF is critical for managing outbreaks and implementing vaccination campaigns. A YF immunoglobulin M (IgM) antibody-capture (MAC) enzyme-linked immunosorbent assay (ELISA) kit, the YF MAC-HD, was successfully introduced starting in 2018 to laboratories in Africa and South America. The YF MAC-HD kit can be performed in 3.5 hours, test up to 24 samples, and includes all reagents necessary to perform the test, except for water used to dilute wash buffer. In 2018 and 2019, a total of 56 laboratory personnel from 39 countries in Africa and South America were trained to use the kit during workshops, followed by take-home YF IgM proficiency testing (PT) exercises. Participants received either a 10- or 20-sample YF PT panel and performed testing using the YF MAC-HD kit. All countries obtained 90% or higher correct results. These results verified the technical viability and transferability of YF MAC-HD kit use for laboratories in YF-endemic countries.

scholarly journals Standardization of Immunoglobulin M Capture Enzyme-Linked Immunosorbent Assays for Routine Diagnosis of Arboviral Infections

2000 ◽  
Vol 38 (5) ◽  
pp. 1823-1826 ◽  
Author(s):  
Denise A. Martin ◽  
David A. Muth ◽  
Teresa Brown ◽  
Alison J. Johnson ◽  
Nick Karabatsos ◽  
...  
Keyword(s):  
Immunoglobulin M ◽  
Detection Sensitivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Virus Infections ◽  
Virus Family ◽  
Serum Dilution ◽  
Routine Diagnosis ◽  
Human Sera ◽  
Antiviral Antibody ◽  
Antibody Capture

Immunoglobulin M antibody-capture enzyme-linked immunosorbent assay (MAC-ELISA) is a rapid and versatile diagnostic method that readily permits the combination of multiple assays. Test consolidation is especially important for arthropod-borne viruses (arboviruses) which belong to at least three virus families: the Togaviridae,Flaviviridae, and Bunyaviridae. Using prototype viruses from each of these families and a panel of well-characterized human sera, we have evaluated and standardized a combined MAC-ELISA capable of identifying virus infections caused by members of each virus family. Furthermore, by grouping antigens geographically and utilizing known serological cross-reactivities, we have reduced the number of antigens necessary for testing, while maintaining adequate detection sensitivity. We have determined that a 1:400 serum dilution is most appropriate for screening antiviral antibody, using a positive-to-negative ratio of ≥2.0 as a positive cutoff value. With a blind-coded human serum panel, this combined MAC-ELISA was shown to have test sensitivity and specificity that correlated well with those of other serological techniques.

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