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      Evaluation of Epic® label-free technology to quantify functional recombinant hemagglutinin

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          Abstract

          Background

          Alternative methods are being sought to measure the potency of influenza vaccines. Label-free technologies that do not require the use of hemagglutinin (HA)-specific antisera are particularly attractive as the preparation of antiserum delays availability of potency reagents. The objective of these experiments was to evaluate the use of a Corning Epic® label-free method to quantify functional influenza hemagglutinin in rHA preparations. The method was optimized to quantify recombinant HA (rHA) of B/Brisbane/60/2008 (B/BR/08). Fetuin was immobilized onto plates and the change in wavelength of refracted light measured using an Enspire (Perkin Elmer) instrument.

          Results

          The change in wavelength measured in response to addition of rHA of B/BR/08 was proportional to its concentration and was optimal in the presence of native rHA conformations. However, the assay was strain-dependent and did not correlate with HAU measured using turkey red blood cells.

          Conclusions

          The Corning Epic® label-free method is suitable for quantifying the native forms of rHA for B/BR/08 and A/Brisbane/59/2007 (H1N1) and A/Hangxhou/3/2013 (H7N9). This method is a useful tool for research purposes but further investigation is needed to identify suitable glycoproteins to use as ligands that allow quantification of HAs from a broader range of virus strains.

          Electronic supplementary material

          The online version of this article (doi:10.1186/s12575-015-0019-5) contains supplementary material, which is available to authorized users.

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          Most cited references20

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          Label-free screening of bio-molecular interactions.

          Lee Cooper (2003)
          The majority of techniques currently employed to interrogate a biomolecular interaction require some type of radio- or enzymatic- or fluorescent-labelling to report the binding event. However, there is an increasing awareness of novel techniques that do not require labelling of the ligand or the receptor, and that allow virtually any complex to be screened with minimal assay development. This review focuses on three major label-free screening platforms: surface plasmon resonance biosensors, acoustic biosensors, and calorimetric biosensors. Scientists in both academia and industry are using biosensors in areas that encompass almost all areas drug discovery, diagnostics, and the life sciences. The capabilities and advantages of each technique are compared and key applications involving small molecules, proteins, oligonucleotides, bacteriophage, viruses, bacteria, and cells are reviewed. The role of the interface between the biosensor surface (in the case of SPR and acoustic biosensors) and the chemical or biological systems to be studied is also covered with attention to the covalent and non-covalent coupling chemistries commonly employed.
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            Label-free cell-based assays with optical biosensors in drug discovery.

            Ye Fang (2006)
            Once viewed solely as a tool for low throughput and kinetic analysis of biomolecular interactions, optical biosensors are gaining widespread uses in drug discovery because of recent advances in instrumentation and experimental design. These advances have expanded the capabilities of optical biosensors to meet the needs at many points in the drug discovery process. Concurrent shifts in drug discovery paradigms have seen the growing use of whole cell systems for drug screens, thus creating both a need in drug discovery and a solution in optical biosensors. This article reviews important advances in optical biosensor instrumentation, and highlights the potential of optical biosensors for drug discovery with an emphasis on whole cell sensing in both high throughput and high content fashions.
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              Properly Folded Bacterially Expressed H1N1 Hemagglutinin Globular Head and Ectodomain Vaccines Protect Ferrets against H1N1 Pandemic Influenza Virus

              Surender Khurana, Swati Verma, Nitin Verma … (2010)
              Background In the face of impending influenza pandemic, a rapid vaccine production and mass vaccination is the most effective approach to prevent the large scale mortality and morbidity that was associated with the 1918 “Spanish Flu”. The traditional process of influenza vaccine production in eggs is time consuming and may not meet the demands of rapid global vaccination required to curtail influenza pandemic. Methodology/Principal Findings Recombinant technology can be used to express the hemagglutinin (HA) of the emerging new influenza strain in a variety of systems including mammalian, insect, and bacterial cells. In this study, two forms of HA proteins derived from the currently circulating novel H1N1 A/California/07/2009 virus, HA1 (1–330) and HA (1–480), were expressed and purified from E. coli under controlled redox refolding conditions that favoured proper protein folding. However, only the recombinant HA1 (1–330) protein formed oligomers, including functional trimers that bound receptor and caused agglutination of human red blood cells. These proteins were used to vaccinate ferrets prior to challenge with the A/California/07/2009 virus. Both proteins induced neutralizing antibodies, and reduced viral loads in nasal washes. However, the HA1 (1–330) protein that had higher content of multimeric forms provided better protection from fever and weight loss at a lower vaccine dose compared with HA (1–480). Protein yield for the HA1 (1–330) ranged around 40 mg/Liter, while the HA (1–480) yield was 0.4–0.8 mg/Liter. Conclusions/Significance This is the first study that describes production in bacterial system of properly folded functional globular HA1 domain trimers, lacking the HA2 transmembrane protein, that elicit potent neutralizing antibody responses following vaccination and protect ferrets from in vivo challenge. The combination of bacterial expression system with established quality control methods could provide a mechanism for rapid large scale production of influenza vaccines in the face of influenza pandemic threat.
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                Author and article information

                Contributors
                Lianlian Jiang: jianglianlian1018@163.com
                Maryna C Eichelberger: Maryna.Eichelberger@fda.hhs.gov
                Journal
                Journal ID (nlm-ta): Biol Proced Online
                Journal ID (iso-abbrev): Biol Proced Online
                Title: Biological Procedures Online
                Publisher: BioMed Central (London )
                ISSN (Print): 1480-9222
                ISSN (Electronic): 1480-9222
                Publication date (Electronic): 9 March 2015
                Publication date PMC-release: 9 March 2015
                Publication date Collection: 2015
                Volume: 17
                Electronic Location Identifier: 7
                Affiliations
                Division of Viral Products, Office of Vaccines Research and Review, Center for Biologics Evaluation and Research, Food and Drug Administration, 10903 New Hampshire Avenue, Silver Spring, MD 20993 USA
                Article
                Publisher ID: 19
                DOI: 10.1186/s12575-015-0019-5
                PMC ID: 4359790
                SO-VID: 052a9580-f46e-40e0-969c-556eb0c85f42
                Copyright © © Jiang and Eichelberger; licensee BioMed Central. 2015
                License:

                This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver ( http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

                History
                Date received : 18 December 2014
                Date accepted : 14 February 2015
                Categories
                Subject: Research
                Custom metadata
                issue-copyright-statement © The Author(s) 2015

                ScienceOpen disciplines: Life sciences
                Keywords: influenza,hemagglutinin,label-free technology,potency,fetuin,corning,epic,enspire,recombinant
                Data availability:
                ScienceOpen disciplines: Life sciences
                Keywords: influenza, hemagglutinin, label-free technology, potency, fetuin, corning, epic, enspire, recombinant

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