Inactivated influenza A/Brisbane/59/2007 H1N1 virus was mixed with PBS and centrifuged at 100.000xg for 1h at 18C. for 30 min). (A) FACS gating for triple co-culture and 16HBecome cells. EpCAM+ cells are 16HBecome cells, EpCAM- cells are further divided into DC-Sign+ (DCs) and DC-Sign- (macrophages) (B) Two times positive CD1c+ and CD11c+ MDDCs were gated according to relevant FMO (fluorescence minus one). Phenotypic and co-stimulatory markers were analyzed according to their relevant isotype settings. (C) Two times positive CD14+ and CD68+ MDMs were gated according to relevant FMO. Phenotypic and AN-3485 co-stimulatory markers were analyzed according to their relevant isotype settings.(EPS) pone.0163539.s002.eps (1.0M) GUID:?8D40C123-8901-4452-9141-8BBBD470ABE9 S3 Fig: Uptake of nanoparticles in EpCAM- mono-culture. Cells were incubated with either virosomes (VIRO), liposomes (LIPO) or control (PBS) for 18h at 37C. Uptake of virosomes and liposomes was determined by measuring Atto647 transmission by circulation cytometry. Rate of recurrence and MFI are demonstrated relative to PBS. Data represents four self-employed experiments. Statistical significance was determined by ANOVA followed by Tukeys HSD post hoc test to investigate individual paired comparisons.(EPS) pone.0163539.s003.eps (1.2M) GUID:?C47A7EA7-8668-463B-Abdominal52-3051A60A9CFB S4 Fig: Manifestation of surface markers in 16HBE cells upon uptake of nanoparticles. Cells in mono- (MO) or co-culture (CO) were incubated for 18h with either virosomes (VIRO), liposomes (LIPO) or settings (PBS, as demonstrated). Manifestation of surface molecule markers HLA-DR, CD40, CD80, CD86 was measured by circulation cytometry. Figures display the receptor manifestation in rate of recurrence of at least six independent experiments. Statistical significance was determined by ANOVA followed by Tukeys HSD post hoc test to investigate individual paired comparisons.(EPS) pone.0163539.s004.eps (1.7M) GUID:?BA4B26A8-C765-48A8-9BEB-CCA78D988544 S5 Fig: Manifestation of surface markers and cytokines in MDDCs upon uptake of nanoparticles in mono-culture. Cells were incubated for 18h with either virosomes (VIRO, with () and without () Atto647), liposomes (LIPO, with () and without () Atto647) or settings (PBS, as demonstrated). Manifestation of surface molecule markers CD83, PD-L1, PD-L2, CCR7 and intracellular cytokines IL-10 and IL-12 was measured by circulation cytometry. Figures display the receptor manifestation in rate of recurrence (A) and MFI (B) of at AN-3485 least six independent experiments. Statistical significance was determined by Mouse monoclonal to CD45 ANOVA followed by Tukeys HSD post hoc test to investigate individual paired comparisons. *p 0.05; **p 0.01; ***p 0.001.(EPS) pone.0163539.s005.eps (2.4M) GUID:?312F4AB8-BF02-4A1D-AAD1-B0220464A149 S6 Fig: Manifestation of cytokines in MDMs upon uptake of nanoparticles in mono-culture. Cells were incubated for 18h with either virosomes (VIRO, with () and without () Atto647), liposomes (LIPO, with () and without () Atto647) or settings (PBS, as demonstrated). Manifestation of intracellular cytokines IL-10 and IL-12 was measured by circulation cytometry. Figures display expression in rate of recurrence (A) and MFI (B) of at least six independent experiments. Statistical significance was determined by ANOVA followed by Tukeys HSD post hoc test to investigate individual paired comparisons. *p 0.05; **p 0.01; ***p 0.001.(EPS) pone.0163539.s006.eps (1.5M) GUID:?906E7E28-CFAF-4AF4-9A72-CDC7861DDFEB S7 Fig: Manifestation of surface markers and cytokines in MDDCs upon treatment with settings in mono-culture. Cells were incubated for 18h with medium (DCs only), or positive settings LPS and inactivated disease A/Brisbane/59/2007 H1N1 (A/B). Manifestation of surface molecule markers HLA-DR, CD40, CD80, CD86, CD83, PD-L1, PD-L2, CCR7 and intracellular cytokines IL-10 and IL-12 was measured by circulation cytometry in MDDCs. Numbers show manifestation in rate of recurrence (A) and MFI (B) of at least six independent experiments. Statistical significance was determined by ANOVA followed by Tukeys HSD post hoc test to investigate individual paired comparisons. *p 0.05; **p 0.01; ***p 0.001.(EPS) pone.0163539.s007.eps (3.3M) GUID:?477681C1-ADD6-4DBE-9E02-8DF855DB0C6D S8 Fig: Manifestation of surface markers and cytokines in MDMs upon treatment with controls in mono-culture. Cells were incubated for 18h with medium (DCs only), or positive settings LPS and inactivated influenza disease A/Brisbane/59/2007 H1N1 (A/B). Manifestation of surface molecule markers HLA-DR, CD40, CD80, CD86, CD163 and intracellular cytokines IL-10 and IL-12 was measured by circulation cytometry. Figures display the receptor manifestation in rate of recurrence (A) and MFI (B) of at least six independent experiments. Statistical significance was determined by ANOVA followed by AN-3485 Tukeys HSD post hoc test to investigate individual paired comparisons. *p 0.05; **p 0.01; ***p 0.001.(EPS) pone.0163539.s008.eps (2.8M) GUID:?A51F7CCD-40F8-4DFF-BAB4-DEED13C166F9 Data Availability StatementAll relevant data are within the paper and its Supporting Info files. Abstract The respiratory tract with its ease of access, vast surface area and dense network of.
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