Journal: Infection & Chemotherapy

Article Title: Humoral and Cellular Immunogenicity Induced by Avian Influenza A (H7N9) DNA Vaccine in Mice

doi: 10.3947/ic.2017.49.2.117

Figure Lengend Snippet: Induction of HA-specific cellular immune responses in mice immunized with H7 HA DNA vaccine . BALB/c mice were immunized with vector (50 µg) or H7 HA DNA vaccine (25 or 50 µg) at one to three doses and spleens were isolated 2 weeks after the last immunization. Mouse splenocytes were stimulated with H7 HA peptide pools for 16 h followed by IFN-γ ELISpot assay. IFN-γ production was determined by calculating the number of spot-forming cells per 1 × 10 6 splenocytes. Naïve mice (N) were used as a negative control. Experiments were performed in triplicate and the values are the means ± standard error of the means. SFU; spot-forming unit.

Article Snippet: Briefly, mice splenocytes were isolated from spleens using a 70-mm nylon cell strainer (BD Pharmingen, San Diego, CA, USA) and incubated in media containing 10 ng/mL interleukin-2 for 48 h. Cells were plated in 96-well multiscreen filter plates (Millipore, Billerica, MA, USA) coated with purified rat anti-mouse IFN-γ capture antibody (BD Pharmingen, San Diego, CA, USA) in triplicate at 1 × 10 6 cells/well and stimulated by H7 HA peptides (GeneOne Life Science, Inc., Seoul, Korea) for 16 h. Biotin rat anti-mouse IFN-γ (BD Pharmingen, San Diego, CA, USA) was used as the detection antibody.

Techniques: Plasmid Preparation, Isolation, Enzyme-linked Immunospot, Negative Control

Journal: Vaccines

Article Title: Increasing the Vaccine Potential of Live M . bovis BCG by Coadministration with Plasmid DNA Encoding a Tuberculosis Prototype Antigen

doi: 10.3390/vaccines2010181

Figure Lengend Snippet: Specific Th1-type cytokine production induced by BCG/pDNA co-vaccination. IFN-γ, IL-2, TNF-α and IL-17A levels (pg/mL) were measured by ELISA in 24 h (IL-2) or 72 h (IFN-γ, TNF-α and IL-17A) culture supernatants of splenocytes from vaccinated C57BL/6 mice, after restimulation with the PPE44 MHC Class II-restricted synthetic peptide spanning amino acids 1–20 (PPE44 [1–20] ) or the recombinant PPE44 protein (rPPE44). RPMI, non-stimulated cells. ID, intradermally. IM, intramuscularly. Data are presented as the mean ± standard error of the mean and representative of two independent experiments, including six mice per group. * p < 0.05; ** p < 0.005.

Article Snippet: IFN-γ was detected using an enzyme-linked immunosorbent assay (ELISA) with purified rat anti-mouse IFN-γ as the capture antibody and biotin-labelled rat anti-mouse IFN-γ as the detection antibody (BD Pharmingen, Franklin Lakes, NJ, USA).

Techniques: Enzyme-linked Immunosorbent Assay, Recombinant

Journal: Vaccines

Article Title: Increasing the Vaccine Potential of Live M . bovis BCG by Coadministration with Plasmid DNA Encoding a Tuberculosis Prototype Antigen

doi: 10.3390/vaccines2010181

Figure Lengend Snippet: Specific CD8 + T-cell responses induced by BCG/pDNA co-vaccination. ( A ) IFN-γ levels (pg/mL) were measured by ELISA in 72 h culture supernatant of splenocytes cultured with the PPE44 MHC Class I-restricted short peptide spanning amino acids 257–265 (PPE44 [257–265] ). RPMI, non-stimulated cells. Data are presented as the mean ± standard error of the mean and representative of two independent experiments, including six mice per group. * p < 0.05. ( B ) In vivo -specific cytotoxic T lymphocyte (CTL) activity. Unpulsed splenocytes (carboxyfluorescein diacetate (CFSE) l ow ) and peptide-pulsed splenocytes (CFSE high ) from naive mice were intravenously transferred to the immunized mice 18 h before flow cytometry analysis. Splenocytes were pulsed with the PPE44 MHC Class I-restrictedshort peptide spanning amino acids 257–265. The percentages of specific lysis in the immunized mice were compared to the ratio in transferred naive control mice. Data represent the mean ± standard error of the mean ( n = 3–6/group). ** p < 0.005. ID, intradermally. IM, intramuscularly.

Article Snippet: IFN-γ was detected using an enzyme-linked immunosorbent assay (ELISA) with purified rat anti-mouse IFN-γ as the capture antibody and biotin-labelled rat anti-mouse IFN-γ as the detection antibody (BD Pharmingen, Franklin Lakes, NJ, USA).

Techniques: Enzyme-linked Immunosorbent Assay, Cell Culture, In Vivo, Activity Assay, Flow Cytometry, Lysis

Journal: Vaccines

Article Title: Increasing the Vaccine Potential of Live M . bovis BCG by Coadministration with Plasmid DNA Encoding a Tuberculosis Prototype Antigen

doi: 10.3390/vaccines2010181

Figure Lengend Snippet: Mycobacteria-specific IFN-γ responses against non plasmid-encoded, but BCG-expressed, antigens after BCG/pDNA co-vaccination. IFN-γ (pg/mL) was measured by ELISA in 72 h culture supernatant of spleen cells cultured with mycobacterial recombinant proteins PstS-3 and Ag85A, BCG culture filtrate (BCG CF) and with recombinant PPE44 ( A ) or purified ovalbumin ( B ). Data are presented as the mean ± standard error of the mean and representative of two independent experiments, including six mice per group. * p < 0.05 in comparison with BCG (ID) and BCG/control pDNA (ID) groups.

Article Snippet: IFN-γ was detected using an enzyme-linked immunosorbent assay (ELISA) with purified rat anti-mouse IFN-γ as the capture antibody and biotin-labelled rat anti-mouse IFN-γ as the detection antibody (BD Pharmingen, Franklin Lakes, NJ, USA).

Techniques: Plasmid Preparation, Enzyme-linked Immunosorbent Assay, Cell Culture, Recombinant, Purification