Fig. 2: Uptake and presentation of the combined LNP vector by APCs in vitro. | Mucosal Immunology

Fig. 2: Uptake and presentation of the combined LNP vector by APCs in vitro.

From: A vaccine combination of lipid nanoparticles and a cholera toxin adjuvant derivative greatly improves lung protection against influenza virus infection

Fig. 2

a M2e-hybridoma T-cell activation by A-20 B cells after incubation with a range of concentrations of non-PEGylated LNPs containing CTA1-3M2e-DD in the inner core (LNP inside), having it covalently attached on the surface (LNP surface) or both (LNP inside + surface). IL-2 production was assessed by proliferation of CTLL-2 cells and given as cell proliferation in c.p.m. (counts per minute) ± SD. The value is the mean of triplicate cultures of each condition in 3 independent experiments. b A similar experimental set up as in (a), but with A-20 B cells incubated with LNPs (inside + outside) of FPM2e, FPM2e:LNP, FPM2e:LNPPEG, or M2e peptide alone. Values are given as mean c.p.m. ± SD of triplicate cultures of each condition in 3 independent experiments. c The uptake, processing and surface presentation of Eα peptide and MHC class II complexes by D1 dendritic cells after incubation for 1 and 4 h with increasing concentrations of Eα-peptide, FPEα, FPEα:LNP, or FPEα:LNPPEG. Surface expression of peptide/MHC II complexes was determined by geometric mean fluorescent intensity (MFI) of labeled Y-Ae Mab by FACS and plotted as mean ± SD of triplicate cultures of each condition in 3 independent experiments. d Same experimental set up as C, but with a fixed protein dose of 0,2 μM of Eα-peptide, FPEα, FPEα:LNP or FPEα:LNPPEG analyzed at different time points. Values are given as mean MFI ± SD of triplicate cultures of each condition in 3 independent experiments (left panel). Representative histograms of Y-Ae MFI for the different conditions after 1 and 4 h of incubation as indicated (right panels). e Using FACS and labeled antibodies we determined the MFI values of MHC II (left panel), CD40 (middle panel) and CD86 (right panel) expression on D1 cells following incubation with LNPs for 1 h, 4 h and 24 h. Values are given as mean MFI ± SD of triplicate cultures of each condition in 3 independent experiments. f The production of IL-1β (left panel), IL-6 (middle panel), and IL-23 (right panel) in culture supernatants from (e) was assessed by ELISA and given as pg/ml ± SD. Statistical significance was calculated by unpaired t test and p values are given as *p < 0.05 and **p < 0.01.

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