Journal: bioRxiv

Article Title: Immunogenicity of an AAV-based, room-temperature stable, single dose COVID-19 vaccine in mouse and non-human primates

doi: 10.1101/2021.01.05.422952

Figure Lengend Snippet: (A) Several RBD-binding antibody isotype titers (IgG, IgG1, IgG2a, IgG2b, IgG3, IgA and IgM) measured weekly in 6-10 week-old BALB/c (n=10, 5 females and 5 males) treated IM with two doses of AC1 and AC3. (B) Ratio of RBD-binding IgG2a and IgG1 antibody titers in serum samples harvested 28 days after vaccination of BALB/c mice as described in A. The Geometric Mean Titer (GMT) is shown above each group. (C and F) Cytokine concentration (pg/mL) in supernatants harvested from splenocytes stimulated for 48h with peptides spanning SARS-CoV-2 Spike protein. Splenocytes were extracted from BALB/c (C) and C57BL/6 (F) animals 4 and 6 weeks, respectively, after vaccination with 10 11 gc of AC1 or AC3. (D-E) Spot forming units (SFU) detected by IFN-γ (D) or IL-4 (E) ELISpot in splenocytes extracted from BALB/c animals 4 weeks after vaccination with 10 11 gc of AC1 or AC3 and stimulated with peptides spanning SARS-CoV-2 Spike protein for 48h. (G-H) Spot forming units (SFU) detected by IFN-γ (G) or IL-4 (H) ELISpot in splenocytes extracted from C57BL/6 animals 6 weeks after vaccination with 10 10 gc of AC1 or AC3 and stimulated with peptides spanning SARS-CoV-2 Spike protein for 48h. For (B-H) data are represented as mean ± SD and groups were compared by Kruskal Wallis and Dunn’s post-test.

Article Snippet: Subsequently, the plates were washed and incubated with biotin-conjugated mouse IFN-γ ELISPOT Detection Antibody (BD Biosciences Cat# 551881, RRID:AB_2868948) and 4 μg/ml biotin-conjugated mouse IL4 detection antibody (BD Biosciences Cat# 551878, RRID:AB_2336921) at room temperature for 3 hours and followed by streptavidin-HRP (dilution 1:1000, Sigma-Aldrich, Cat# 18-152) for 45 minutes.

Techniques: Binding Assay, Concentration Assay, Enzyme-linked Immunospot

Journal: Frontiers in Immunology

Article Title: Interleukin-33 Contributes to the Induction of Th9 Cells and Antitumor Efficacy by Dectin-1-Activated Dendritic Cells

doi: 10.3389/fimmu.2018.01787

Figure Lengend Snippet: Interleukin (IL)-33/ST2 axis contributes to dectin-1-activated dendritic cell (DC)-induced Th9 priming in vivo . (A,B) OT-II mice (two to three mice/group) were immunized twice (1 week apart) with OVA-peptide-pulsed BMDCs or CurDCs. PBS served as a control. On day 3 after the second immunization, spleen cells were harvested and restimulated separately with OVA-peptide-pulsed BMDCs and CurDCs for 2 days in the culture. (A) Flow cytometry of ST2-expressing CD4 + T cells. Numbers in the dot plots represent the percentages of ST2 + CD4 + T cells. Right, summarized results of three independent experiments obtained as at left. The experiments were performed three times ( n = 3). (B) Quantitative polymerase chain reaction (qPCR) analysis of St2 in CD4 + T cells. The experiments were performed three times ( n = 3). (C,D) OT-II mice (two to three mice/group) were immunized twice (1 week apart) with OVA-peptide-pulsed CurDCs in the presence of control IgG or αST2. PBS served as a control. On day 3 after the second immunization, spleen cells were restimulated separately with OVA-peptide-pulsed CurDCs plus IgG or αST2 for 48 h. (C) qPCR analyses of Il9, Ifng , and Gzmb in CD4 + T cells. The experiments were performed three times ( n = 3). (D) Enzyme-linked immunosorbent assay (ELISA) assessed IL-9 and IFN-γ in the cultures. The experiments were performed three times ( n = 3). (E–H) OT-II mice (two to three mice/group) were immunized twice (1 week apart) with OVA-peptide-pulsed BMDCs or CurDCs. In addition, some of the mice immunized with BMDCs or CurDCs were given IL-33 (250 ng/mouse) every 3 days, beginning on the day of the first immunization. Mice received PBS served as controls. Mouse spleen cells were restimulated with BMDCs or CurDCs in the presence or absence of IL-33 (50 ng/mL) for 2 days in the culture. (E) Flow cytometry of ST2-, IFN-γ-, IL-9-, or GzmB-producing CD4 + T cells. Numbers in the dot plots represent the percentages of double-positive T helper cells. (F) Summarized results of three independent experiments obtained in (A) . The experiments were performed three times ( n = 3). (G) ELISA assays of IL-9 and IFN-γ in the culture. The experiments were performed three times ( n = 3). (H) qPCR analyses of St2, Il9, Ifng , and Gzmb in CD4 + T cells. The experiments were performed three times ( n = 3). Data are representative of three (A,E) independent experiments or presented as mean ± SD of three (A–D,F–H) independent experiments. NS , non-significant; * P < 0.05; ** P < 0.01.

Article Snippet: Capture/detection Abs for IL-9 and IFN-γ were purchased from BD Biosciences.

Techniques: In Vivo, Flow Cytometry, Expressing, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

Journal: BMC Infectious Diseases

Article Title: Potential of novel Mycobacterium tuberculosis infection phase-dependent antigens in the diagnosis of TB disease in a high burden setting

doi: 10.1186/1471-2334-12-10

Figure Lengend Snippet: IFN-γ levels (pg/ml) elicited by classical TB antigens in 23 TB patients (TB) and 20 household contacts (HHC) . Responses in TB cases are indicated by open symbols and those in household contacts by closed symbols. Error bars represent the median. Ag85A/B = Ag85A and Ag85B tested together, HSP = heat shock protein, PHA = Phytohaemagglutinin.

Article Snippet: Plates were then washed and 1 μg/ml (100 μl) of the biotinylated mouse-anti-human IFN-γ detection antibody (BD PharmingenTM) added.

Techniques:

Journal: BMC Infectious Diseases

Article Title: Potential of novel Mycobacterium tuberculosis infection phase-dependent antigens in the diagnosis of TB disease in a high burden setting

doi: 10.1186/1471-2334-12-10

Figure Lengend Snippet: Number of inclusions of antigens into the 10 general discriminant analysis models that most accurately predicted the presence or absence of TB disease . PHA was evaluated in all study participants as a positive control, and appeared in one of the models because IFN-γ data was analyzed in a blinded manner.

Article Snippet: Plates were then washed and 1 μg/ml (100 μl) of the biotinylated mouse-anti-human IFN-γ detection antibody (BD PharmingenTM) added.

Techniques: Positive Control