Use of an Influenza Antigen Microarray to Measure the Breadth of Serum Antibodies Across Virus Subtypes
Autor: | Li Liang, Jiin Felgner, Erwin Strahsburger, D. Huw Davies, Rie Nakajima, Jenny Davies, Philip L. Felgner, Algis Jasinskas, Egest J. Pone, Saahir Khan, Rafael Ramiro de Assis, Omid Taghavian, Aarti Jain, Sharon Jan, Medalyn Supnet, Joshua M. Obiero |
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Rok vydání: | 2019 |
Předmět: |
0301 basic medicine
General Chemical Engineering Protein Array Analysis Hemagglutinin (influenza) Neuraminidase Enzyme-Linked Immunosorbent Assay Hemagglutinin Glycoproteins Influenza Virus 02 engineering and technology Cross Reactions Antibodies Viral Virus Antigenic drift General Biochemistry Genetics and Molecular Biology Cohort Studies 03 medical and health sciences Viral Proteins Antigen Influenza Human Humans Prospective Studies Antigens Viral biology General Immunology and Microbiology General Neuroscience Reproducibility of Results 021001 nanoscience & nanotechnology Virology Primary and secondary antibodies Antibodies Neutralizing 030104 developmental biology Influenza Vaccines biology.protein Protein microarray Antibody 0210 nano-technology |
Zdroj: | Journal of Visualized Experiments. |
ISSN: | 1940-087X |
Popis: | The influenza virus remains a significant cause of mortality worldwide due to the limited effectiveness of currently available vaccines. A key challenge to the development of universal influenza vaccines is high antigenic diversity resulting from antigenic drift. Overcoming this challenge requires novel research tools to measure the breadth of serum antibodies directed against many virus strains across different antigenic subtypes. Here, we present a protocol for analyzing the breadth of serum antibodies against diverse influenza virus strains using a protein microarray of influenza antigens. This influenza antigen microarray is constructed by printing purified hemagglutinin and neuraminidase antigens onto a nitrocellulose-coated membrane using a microarray printer. Human sera are incubated on the microarray to bind antibodies against the influenza antigens. Quantum-dot-conjugated secondary antibodies are used to simultaneously detect IgG and IgA antibodies binding to each antigen on the microarray. Quantitative antibody binding is measured as fluorescence intensity using a portable imager. Representative results are shown to demonstrate assay reproducibility in measuring subtype-specific and cross-reactive influenza antibodies in human sera. Compared to traditional methods such as ELISA, the influenza antigen microarray provides a high throughput multiplexed approach capable of testing hundreds of sera for multiple antibody isotypes against hundreds of antigens in a short time frame, and thus has applications in sero-surveillance and vaccine development. A limitation is the inability to distinguish binding antibodies from neutralizing antibodies. |
Databáze: | OpenAIRE |
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