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Development of a duplex Fluorescent Microsphere Immunoassay (FMIA) for the detection of antibody responses to influenza A and newcastle disease viruses
- Source :
- Journal of Immunological Methods. 405:167-177
- Publication Year :
- 2014
- Publisher :
- Elsevier BV, 2014.
-
Abstract
- Highly pathogenic avian influenza virus (HPAI) and virulent forms of avian paramyxovirus-1 (APMV-1) cause serious illnesses in domestic poultry, both of which are reportable to the World Organization of Animal Health (OIE). The clinical presentation of avian influenza (AI) and APMV-1 infections are difficult to differentiate, emphasizing the importance of rapid and sensitive serologic assays that are able to distinguish them. Currently, a variety of serological assays are used for the serologic diagnosis of both diseases, but these assays are not used in multiplex formats. In this study, development of a duplex fluorescent microsphere immunoassay (FMIA) based on Luminex xMAP Technology is described. The assay employs MagPlex magnetic microspheres that are covalently coated with recombinant avian influenza virus nucleoprotein and APMV-1 nucleocapsid antigens produced in a baculovirus insect cell expression system. The assay is able to detect AIV antibodies against all existing hemagglutinin (H1–H16) subtypes and simultaneously detect antibodies against APMV-1. In the process of this assay development different bead coupling conditions were compared. The assay has the capability of detecting serum antibodies from chickens and turkeys and optimization was accomplished by using 2462 chicken and 446 turkey field and experimental sera and had a comparable detection capability with currently used assays in the laboratory. Assay threshold values were calculated with Receiver Operating Characteristic Analysis (ROC) in non-parametric analysis due to a highly skewed data distribution; this analysis resulted in AIV nucleoprotein relative diagnostic sensitivity and specificity of 99.7%, and 97.3% respectively. The APMV-1 nucleocapsid relative diagnostic sensitivity and specificity were 95.4%, and 98.5% respectively.
- Subjects :
- Turkeys
Newcastle Disease
Blotting, Western
Immunology
Newcastle disease virus
APMV-1
Spodoptera
Biology
Antibodies, Viral
medicine.disease_cause
Newcastle disease
Fluorescence
Serology
Antigen
Sf9 Cells
Luminex
Influenza A virus
medicine
Animals
Immunology and Allergy
Multiplex
FMIA
Immunoassay
medicine.diagnostic_test
Reproducibility of Results
Nucleocapsid Proteins
biology.organism_classification
Virology
Microspheres
Recombinant Proteins
Influenza A virus subtype H5N1
Nucleoprotein
ROC Curve
Influenza in Birds
Antibody Formation
Chickens
Avian Influenza
Subjects
Details
- ISSN :
- 00221759
- Volume :
- 405
- Database :
- OpenAIRE
- Journal :
- Journal of Immunological Methods
- Accession number :
- edsair.doi.dedup.....a4ed4974fd0e8fde2ea2facc0e6297fc