1. Production and application of anti-nucleoprotein IgY antibodies for influenza A virus detection in swine.
- Author
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da Silva MC, Schaefer R, Gava D, Souza CK, da Silva Vaz I Jr, Bastos AP, and Venancio EJ
- Subjects
- Animals, Chickens immunology, Dogs, Enzyme-Linked Immunosorbent Assay methods, Humans, Madin Darby Canine Kidney Cells, Swine, Antibodies, Viral chemistry, Antibodies, Viral immunology, Immunoglobulins chemistry, Immunoglobulins immunology, Influenza A virus immunology, Influenza A virus metabolism, Orthomyxoviridae Infections blood, Orthomyxoviridae Infections immunology, Orthomyxoviridae Infections veterinary, Swine Diseases blood, Swine Diseases immunology, Swine Diseases virology, Viral Core Proteins blood, Viral Core Proteins immunology
- Abstract
Influenza A virus (IAV) causes an important respiratory disease in mammals and birds leading to concerns in animal production industry and public health. Usually, antibodies produced in mammals are employed in diagnostic tests. However, due to animal welfare concerns, technical advantages and the high cost of production, alternatives to the production of antibodies in mammals have been investigated. The aim of this study was to produce egg yolk immunoglobulin (IgY) in laying hens against a highly conserved protein (nucleoprotein- NP) of IAV and to evaluate the application of anti-NP IgY antibodies in virus detection by immunocytochemistry (ICC) and immunohistochemistry (IHC). Three laying hens of the White Leghorn line were inoculated seven times with a recombinant NP protein and their eggs collected seven days after the 3rd, 5th and 7th inoculations. Immunoglobulin Y antibodies were purified from egg yolk through precipitation with ammonium sulfate. The titers and specificity of the purified antibodies were determined by ELISA, western blotting, ICC and IHC. High levels of specific anti-NP antibodies were detected by ELISA after the 5th inoculation, reaching a peak after the 7th inoculation. The mean yield of total protein in yolk after the 7th inoculation was 13.5 mg/mL. The use of western blotting and ICC demonstrated that anti-NP IgY binds specifically to NP protein. Moreover, the use of anti-NP IgY antibody in ICC test revealed positive staining of MDCK cells infected with IAV of the three subtypes circulating in swine (H1N1, H1N2, and H3N2). However, no staining was observed in lung tissues through the IHC test. The data obtained showed that anti-NP IgY antibodies bound specifically to influenza virus NP protein, detecting the main virus subtypes circulating in swine, reinforcing their usefulness in the influenza diagnosis., (Copyright © 2018 Elsevier B.V. All rights reserved.)
- Published
- 2018
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