tetano
Editor, Senior Moderator
FASEB J. 2015 Jun 2. pii: fj.15-270421. [Epub ahead of print]
[h=1]Generation and characterization of a trackable plant-made influenza H5 virus-like particle (VLP) containing enhanced green fluorescent protein (eGFP).[/h] Young KR[SUP]1[/SUP], Arthus-Cartier G[SUP]1[/SUP], Yam KK[SUP]1[/SUP], Lavoie PO[SUP]1[/SUP], Landry N[SUP]1[/SUP], D'Aoust MA[SUP]1[/SUP], V?zina LP[SUP]1[/SUP], Couture MM[SUP]1[/SUP], Ward BJ[SUP]2[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] Medicago, Inc. has developed an efficient virus-like particle (VLP) vaccine production platform using the Nicotiana benthamiana expression system, and currently has influenza-based products targeting seasonal/pandemic hemagglutinin (HA) proteins in advanced clinical trials. We wished to generate a trackable HA-based VLP that would allow us to study both particle assembly in plants and VLP interactions within the mammalian immune system. To this end, a fusion protein was designed, composed of H5 (from influenza A/Indonesia/05/2005 [H5N1]) with enhanced green fluorescent protein (eGFP). Expression of H5-eGFP in N. benthamiana produced brightly fluorescent ∼160 nm particles resembling H5-VLPs. H5-eGFP-VLPs elicited anti-H5 serologic responses in mice comparable to those elicited by H5-VLPs in almost all assays tested (hemagglutination inhibition/IgG(total)/IgG1/IgG2b/IgG2a:IgG1 ratio), as well as a superior anti-GFP IgG response (mean optical density = 2.52 ? 0.16 sem) to that elicited by soluble GFP (mean optical density = 0.12 ? 0.06 sem). Confocal imaging of N. benthamiana cells expressing H5-eGFP displayed large fluorescent accumulations at the cell periphery, and draining lymph nodes from mice given H5-eGFP-VLPs via footpad injection demonstrated bright fluorescence shortly after administration (10 min), providing proof of concept that the H5-eGFP-protein/VLPs could be used to monitor both VLP assembly and immune trafficking. Given these findings, this novel fluorescent reagent will be a powerful tool to gain further fundamental insight into the biology of influenza VLP vaccines.-Young, K. R., Arthus-Cartier, G., Yam, K. K., Lavoie, P.-O., Landry, N., D'Aoust, M.-A., V?zina, L.-P., Couture, M. M.-J., Ward, B. J. Generation and characterization of a trackable plant-made influenza H5 virus-like particle (VLP) containing enhanced green fluorescent protein (eGFP).
? FASEB.
[h=4]KEYWORDS:[/h] H5 hemagglutinin; fluorescent particle; immune response; particle assembly
PMID: 26038124 [PubMed - as supplied by publisher]
[h=1]Generation and characterization of a trackable plant-made influenza H5 virus-like particle (VLP) containing enhanced green fluorescent protein (eGFP).[/h] Young KR[SUP]1[/SUP], Arthus-Cartier G[SUP]1[/SUP], Yam KK[SUP]1[/SUP], Lavoie PO[SUP]1[/SUP], Landry N[SUP]1[/SUP], D'Aoust MA[SUP]1[/SUP], V?zina LP[SUP]1[/SUP], Couture MM[SUP]1[/SUP], Ward BJ[SUP]2[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] Medicago, Inc. has developed an efficient virus-like particle (VLP) vaccine production platform using the Nicotiana benthamiana expression system, and currently has influenza-based products targeting seasonal/pandemic hemagglutinin (HA) proteins in advanced clinical trials. We wished to generate a trackable HA-based VLP that would allow us to study both particle assembly in plants and VLP interactions within the mammalian immune system. To this end, a fusion protein was designed, composed of H5 (from influenza A/Indonesia/05/2005 [H5N1]) with enhanced green fluorescent protein (eGFP). Expression of H5-eGFP in N. benthamiana produced brightly fluorescent ∼160 nm particles resembling H5-VLPs. H5-eGFP-VLPs elicited anti-H5 serologic responses in mice comparable to those elicited by H5-VLPs in almost all assays tested (hemagglutination inhibition/IgG(total)/IgG1/IgG2b/IgG2a:IgG1 ratio), as well as a superior anti-GFP IgG response (mean optical density = 2.52 ? 0.16 sem) to that elicited by soluble GFP (mean optical density = 0.12 ? 0.06 sem). Confocal imaging of N. benthamiana cells expressing H5-eGFP displayed large fluorescent accumulations at the cell periphery, and draining lymph nodes from mice given H5-eGFP-VLPs via footpad injection demonstrated bright fluorescence shortly after administration (10 min), providing proof of concept that the H5-eGFP-protein/VLPs could be used to monitor both VLP assembly and immune trafficking. Given these findings, this novel fluorescent reagent will be a powerful tool to gain further fundamental insight into the biology of influenza VLP vaccines.-Young, K. R., Arthus-Cartier, G., Yam, K. K., Lavoie, P.-O., Landry, N., D'Aoust, M.-A., V?zina, L.-P., Couture, M. M.-J., Ward, B. J. Generation and characterization of a trackable plant-made influenza H5 virus-like particle (VLP) containing enhanced green fluorescent protein (eGFP).
? FASEB.
[h=4]KEYWORDS:[/h] H5 hemagglutinin; fluorescent particle; immune response; particle assembly
PMID: 26038124 [PubMed - as supplied by publisher]