Journal: Immunology letters

Article Title: A lack of confirmation with alternative assays questions the validity of IL-17A expression in human neutrophils using immunohistochemistry.

doi: 10.1016/j.imlet.2014.10.025

Figure Lengend Snippet: Fig. 1. Detection of IL-17A in human neutrophils by IHC. (A) Cyanine3 labelled IL-17A+ cells infiltrate Wolbachia-containing nodules but not Wolbachia-depleted nodules. (B) Counterstaining

Article Snippet: Immunoprecipitation was carried out using 100 g mouse anti-human IL-17A antibody (41802, R&D Systems) and 60 g goat anti-human IL-17A antibody (AF-317-NA, R&D Systems) coupled to 1 mL HiTrap NHS-activated HP columns and AKTAprime Plus low-pressure affinity chromatography system (both from GE Healthcare) using the manufacturer’s protocol.

Techniques:

Journal: Immunology letters

Article Title: A lack of confirmation with alternative assays questions the validity of IL-17A expression in human neutrophils using immunohistochemistry.

doi: 10.1016/j.imlet.2014.10.025

Figure Lengend Snippet: Fig. 2. Determination of IL-17A expression by human neutrophils. (A) Western blotting of neutrophil (PMN) cell lysate using goat anti-IL-17A antibody and mouse anti- IL-17A antibody.

Article Snippet: Immunoprecipitation was carried out using 100 g mouse anti-human IL-17A antibody (41802, R&D Systems) and 60 g goat anti-human IL-17A antibody (AF-317-NA, R&D Systems) coupled to 1 mL HiTrap NHS-activated HP columns and AKTAprime Plus low-pressure affinity chromatography system (both from GE Healthcare) using the manufacturer’s protocol.

Techniques: Expressing, Western Blot

Journal: Allergology international : official journal of the Japanese Society of Allergology

Article Title: Interleukin-17A expression in human synovial mast cells in rheumatoid arthritis and osteoarthritis.

doi: 10.1016/j.alit.2016.04.007

Figure Lengend Snippet: Fig. 1. Expression of IL-17A in synovial MCs from OA patients and from RA patients. Representative images of IL-17Aþ MCs (tryptaseþ cells) in synovia from an OA patient (A) and an RA patient (B). Immunochemical staining of synovia with anti-tryptase mAb (green, a) and anti-IL-17A (red, b) Ab, and a merged image (yellow, c). The cells were stained with DAPI (blue). The white arrow heads indicate tryptase-IL-17A double-positive cells. Bar ¼ 10 mm. (C) As negative controls, a synovium sample from an OA patient was stained with mouse IgG1 and goat IgG. The results are representative of separate analyses using 6 patients with OA and 6 patients with RA.

Article Snippet: The following antibodies (Abs) were purchased from the indicated sources: human IgE (Calbiochem, San Diego, CA, USA); goat anti-human IL-17A polyclonal Ab (R&D Systems, Minneapolis, MN, USA); anti-human tryptase monoclonal (m) Ab (clone AA1; DakoCytomation Inc., Carpinteria, CA, USA); anti-Fc 3RIa mAb (clone CRA1; eBioscience, San Diego, CA, USA); anti-human IgE Ab (DakoCytomation Inc.); and F(ab0)2 fragments of anti-human FcgRI (clone 10.1; ID Labs Inc., London, ON, Canada).

Techniques: Expressing, Staining

Journal: Allergology international : official journal of the Japanese Society of Allergology

Article Title: Interleukin-17A expression in human synovial mast cells in rheumatoid arthritis and osteoarthritis.

doi: 10.1016/j.alit.2016.04.007

Figure Lengend Snippet: Fig. 2. A comparison of the numbers of MCs (A), the numbers of IL-17Aþ MCs (B), the numbers of IL-17Aþ cells (C), the percentage of IL-17Aþ MCs among all the MCs (D), and the percentage of IL-17Aþ MCs among all the IL-17Aþ cells (E) in synovia from OA patients (open circles, n ¼ 6) and from RA patients (closed circles, n ¼ 6). Data are expressed as the median and inter-quartile range. N.S., not significant.

Article Snippet: The following antibodies (Abs) were purchased from the indicated sources: human IgE (Calbiochem, San Diego, CA, USA); goat anti-human IL-17A polyclonal Ab (R&D Systems, Minneapolis, MN, USA); anti-human tryptase monoclonal (m) Ab (clone AA1; DakoCytomation Inc., Carpinteria, CA, USA); anti-Fc 3RIa mAb (clone CRA1; eBioscience, San Diego, CA, USA); anti-human IgE Ab (DakoCytomation Inc.); and F(ab0)2 fragments of anti-human FcgRI (clone 10.1; ID Labs Inc., London, ON, Canada).

Techniques: Comparison

Journal: Allergology international : official journal of the Japanese Society of Allergology

Article Title: Interleukin-17A expression in human synovial mast cells in rheumatoid arthritis and osteoarthritis.

doi: 10.1016/j.alit.2016.04.007

Figure Lengend Snippet: Fig. 3. IL-17A production from cultured synovium-derived MC following IgE- and IgG-dependent stimulation. (A) IgE-sensitized MCs from patients with OA (white bars, n ¼ 4 donors) or from patients with RA (black bars, n ¼ 4 donors) were incubated with anti-IgE for 6 h. (B) MCs from patients with OA (n ¼ 4 donors) were incubated with 1 mg/ml of F(ab0)2aFcgRI (black bars) or F(ab0)2mIgG1 (white bars) for 30 min and were then stimulated with gF(ab0)2amF(ab0)2 for 6 h. (C) MCs from patients with OA (n ¼ 6 donors) were stimulated with monomeric IgG or aggregated IgG for 6 h. (D) MCs from patients with OA (white bars, n ¼ 7 donors) and from patients with RA (black bars, n ¼ 4 donors) were stimulated with IL-33 for 6 h. (E, F) MCs from patients with OA (n ¼ 5e6 donors) were stimulated with anti-Fc

Article Snippet: The following antibodies (Abs) were purchased from the indicated sources: human IgE (Calbiochem, San Diego, CA, USA); goat anti-human IL-17A polyclonal Ab (R&D Systems, Minneapolis, MN, USA); anti-human tryptase monoclonal (m) Ab (clone AA1; DakoCytomation Inc., Carpinteria, CA, USA); anti-Fc 3RIa mAb (clone CRA1; eBioscience, San Diego, CA, USA); anti-human IgE Ab (DakoCytomation Inc.); and F(ab0)2 fragments of anti-human FcgRI (clone 10.1; ID Labs Inc., London, ON, Canada).

Techniques: Cell Culture, Derivative Assay, Incubation

Journal: Allergology international : official journal of the Japanese Society of Allergology

Article Title: Interleukin-17A expression in human synovial mast cells in rheumatoid arthritis and osteoarthritis.

doi: 10.1016/j.alit.2016.04.007

Figure Lengend Snippet: Fig. 4. IL-17A (A) and IL-8 production (B) from cultured synovium-derived MC following various stimulations. MCs from patients with OA (n ¼ 3 donors) were stimulated with TNF- a, C5a, LPS, or IL-23 plus IL-1b for 24 h. IL-17 levels were under the detection limit in 2 donors out of 3 donors. The data of IL-17A production from one donor are shown. The data of IL-8 production are shown as the mean ± SEM. *P < 0.05.

Article Snippet: The following antibodies (Abs) were purchased from the indicated sources: human IgE (Calbiochem, San Diego, CA, USA); goat anti-human IL-17A polyclonal Ab (R&D Systems, Minneapolis, MN, USA); anti-human tryptase monoclonal (m) Ab (clone AA1; DakoCytomation Inc., Carpinteria, CA, USA); anti-Fc 3RIa mAb (clone CRA1; eBioscience, San Diego, CA, USA); anti-human IgE Ab (DakoCytomation Inc.); and F(ab0)2 fragments of anti-human FcgRI (clone 10.1; ID Labs Inc., London, ON, Canada).

Techniques: Cell Culture, Derivative Assay

Journal: Frontiers in Immunology

Article Title: Age-related increase of mitochondrial content in human memory CD4+ T cells contributes to ROS-mediated increased expression of proinflammatory cytokines

doi: 10.3389/fimmu.2022.911050

Figure Lengend Snippet: Capacity of cytokine production in memory CD4 + T cell. Cells were activated with PMA/ionomycin for 5 h, adding BFA for the last 3 hours for intracellular cytokine expression. (A) Representative example of cytokine expression in memory CD4 + T cells from young and aged donors after stimulation with PMA/ionomycin. (B) Frequencies in % and geometric mean fluorescence intensity (gMFI) of IL-2, IL-4, IL-10, IL-17, IFN-γ, and TNF-α producing memory CD4 + T cells (mean ± SEM; n = 7).

Article Snippet: Incubation was conducted by dissolving 1x 10 6 cells/ml in RPMI 1640 medium (Thermo Fisher Scientific) with 5% human AB serum (Sigma Aldrich) and 1% penicillin/streptomycin (Thermo Fisher Scientific) at 37˚C in a 5% CO 2 atmosphere Cells were collected, washed, fixed, permeabilized (Inside staining kit, Miltenyi Biotec) and stained with a combination of monoclonal antibodies including anti-human-IFNγ (PERCP-Cy5.5, Biolegend, clone 4SB3, Cat.# 502526), anti-human-IL-2 (APC-vio770, Miltenyi Biotec, clone N7.48 A, Cat.# 130-097-011), anti-human-IL-4 (PE, Miltenyi Biotec, clone 7A3-3, Cat.# 130-091-647), anti-human-IL-17A (FITC, Miltenyi Biotec, clone CZ8-23G1, Cat.# 130-094-520), and anti-human-TNF-α (PE-Vio770 (Miltenyi Biotec, clone cA2, Cat.# 130-096-755).

Techniques: Expressing, Fluorescence