Polyclonal antibody production for membrane proteins via genetic immunization

Debra T. Hansen, Mark D. Robida, Felicia M. Craciunescu, Andrey V. Loskutov, Katerina Dörner, John Charles Rodenberry, Xiao Wang, Tien L. Olson, Hetal Patel, Petra Fromme, Kathryn F. Sykes

Research output: Contribution to journalArticle

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Abstract

Antibodies are essential for structural determinations and functional studies of membrane proteins, but antibody generation is limited by the availability of properly-folded and purified antigen. We describe the first application of genetic immunization to a structurally diverse set of membrane proteins to show that immunization of mice with DNA alone produced antibodies against 71% (n = 17) of the bacterial and viral targets. Antibody production correlated with prior reports of target immunogenicity in host organisms, underscoring the efficiency of this DNA-gold micronanoplex approach. To generate each antigen for antibody characterization, we also developed a simple in vitro membrane protein expression and capture method. Antibody specificity was demonstrated upon identifying, for the first time, membrane-directed heterologous expression of the native sequences of the FopA and FTT1525 virulence determinants from the select agent Francisella tularensis SCHU S4. These approaches will accelerate future structural and functional investigations of therapeutically-relevant membrane proteins.

Original languageEnglish
Article number21925
JournalScientific Reports
Volume6
DOIs
Publication statusPublished - Feb 24 2016

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Antibody Formation
Immunization
Membrane Proteins
Antibodies
Francisella tularensis
Antigens
Antibody Specificity
DNA
Gold
Virulence
Membranes

ASJC Scopus subject areas

  • General

Cite this

Hansen, D. T., Robida, M. D., Craciunescu, F. M., Loskutov, A. V., Dörner, K., Rodenberry, J. C., ... Sykes, K. F. (2016). Polyclonal antibody production for membrane proteins via genetic immunization. Scientific Reports, 6, [21925]. https://doi.org/10.1038/srep21925

Polyclonal antibody production for membrane proteins via genetic immunization. / Hansen, Debra T.; Robida, Mark D.; Craciunescu, Felicia M.; Loskutov, Andrey V.; Dörner, Katerina; Rodenberry, John Charles; Wang, Xiao; Olson, Tien L.; Patel, Hetal; Fromme, Petra; Sykes, Kathryn F.

In: Scientific Reports, Vol. 6, 21925, 24.02.2016.

Research output: Contribution to journalArticle

Hansen, DT, Robida, MD, Craciunescu, FM, Loskutov, AV, Dörner, K, Rodenberry, JC, Wang, X, Olson, TL, Patel, H, Fromme, P & Sykes, KF 2016, 'Polyclonal antibody production for membrane proteins via genetic immunization', Scientific Reports, vol. 6, 21925. https://doi.org/10.1038/srep21925
Hansen DT, Robida MD, Craciunescu FM, Loskutov AV, Dörner K, Rodenberry JC et al. Polyclonal antibody production for membrane proteins via genetic immunization. Scientific Reports. 2016 Feb 24;6. 21925. https://doi.org/10.1038/srep21925
Hansen, Debra T. ; Robida, Mark D. ; Craciunescu, Felicia M. ; Loskutov, Andrey V. ; Dörner, Katerina ; Rodenberry, John Charles ; Wang, Xiao ; Olson, Tien L. ; Patel, Hetal ; Fromme, Petra ; Sykes, Kathryn F. / Polyclonal antibody production for membrane proteins via genetic immunization. In: Scientific Reports. 2016 ; Vol. 6.
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