Journal: Bioinformatics advances

Article Title: Different approaches to Imaging Mass Cytometry data analysis.

doi: 10.1093/bioadv/vbad046

Figure Lengend Snippet: Fig. 2. A schematic overview of the Imaging Mass Cytometry workflow (created with BioRender.com)

Article Snippet: IMC was developed in 2014 based on earlier available suspensionbased, single-cell mass cytometry [cytometry time of flight (CyTOF)] technology [described in Bandura et al. (2009)] but combined with an additional platform for UV ablation (Hyperion Tissue Imager, Standard BioTools, South San Francisco, USA) and performed on stained tissue sections, giving the spatial resolution of the data (Fig. 1A) (Bandura et al., 2009).

Techniques: Imaging, Mass Cytometry

Journal: Journal for Immunotherapy of Cancer

Article Title: IL-31 induces antitumor immunity in breast carcinoma

doi: 10.1136/jitc-2020-001010

Figure Lengend Snippet: IL-31 expression in tumors enhances antitumor immunity. (A) Non-tumor-bearing mice were infused with IL-31 via osmotic pumps (~14 µg/day) or control (n=5 mice/group). Three weeks later, mice were sacrificed and spleens were removed. Splenocytes (pooled per group) were immunostained with a 37-antibody panel as described in . Cells were acquired by CyTOF and data were analyzed by Cytobank. The analysis of the different immune cell populations found in the spleens is presented by viSNE plot. (B–D) PyMT cells stably overexpressing IL-31 (PyMT-IL-31) or their counterpart control cells (PyMT-ev) were implanted in the mammary fat pads of 10-week-old C57BL/6 female mice (n=6 mice/group). Tumor volume was measured and plotted (B). At endpoint, tumors were removed and prepared as single cell suspensions. Lymphoid cells (C), Granzyme B expression in tumor lysates (D), and myeloid cells (E) were assessed. Statistical significance was assessed by unpaired two-tailed t-test. Significant p values are shown. CTL, cytotoxic T lymphocyte; CyTOF, time of flight mass cytometry; IL-31, interleukin-31; MDSCs, myeloid-derived suppressor cells; NK, natural killer; ns, non-significant; PMN, polymorphonuclear; TAM, tumor-associated macrophage; Th, T helper.

Article Snippet: The cells were acquired by time of flight mass cytometry (CyTOF) (Fluidigm, South San Francisco, CA, USA) as previously described.

Techniques: Expressing, Control, Stable Transfection, Two Tailed Test, Mass Cytometry, Derivative Assay

Journal: Journal for Immunotherapy of Cancer

Article Title: IL-31 induces antitumor immunity in breast carcinoma

doi: 10.1136/jitc-2020-001010

Figure Lengend Snippet: IL-31 promotes a cytotoxic T cell memory phenotype and inhibits MDSC motility. (A) CD8 + T cells isolated from naïve mice were co-cultured with Gr1 + MDSCs isolated from PyMT tumors. T cells were activated with soluble anti-CD3 and anti-CD28. Cultures were then supplemented with 100 ng/mL IL-31 or left untreated. After 4 days, the activation and memory phenotype of CD8 + T cells were evaluated by flow cytometry. (B) The expression of CD44 and CD62L was evaluated in PMN-MDSCs isolated from peripheral blood of mice implanted with PyMT-ev or PyMT-IL-31 tumors. The evaluation was performed by flow cytometry using MFI. (C, D) The motility of MDSCs was measured by seeding one million splenocytes from naïve C57BL/6 mice (n=4 biological repeats) on a 3 µm insert containing SF media in both upper and lower compartments. The cells were treated with 100 ng/mL IL-31 or left untreated (control). After 18 hours, the cells in the lower compartment were collected, stained for Ly6G and Ly6C, and counted by flow cytometry. A summary graph is shown (C) followed by representative flow cytometry dot-plots (D). Statistical significance was assessed by unpaired two-tailed t-test. Statistical significance of MDSC motility was assessed by paired t-test. Significant p values are shown. CTL, cytotoxic T lymphocytes; IL-31, interleukin-31; MDSCs, myeloid-derived suppressor cells; MFI, mean fluorescence intensity; PMN, polymorphonuclear; SF, serum free media.

Article Snippet: The cells were acquired by time of flight mass cytometry (CyTOF) (Fluidigm, South San Francisco, CA, USA) as previously described.

Techniques: Isolation, Cell Culture, Activation Assay, Flow Cytometry, Expressing, Control, Staining, Two Tailed Test, Derivative Assay, Fluorescence

Journal: Journal for Immunotherapy of Cancer

Article Title: IL-31 induces antitumor immunity in breast carcinoma

doi: 10.1136/jitc-2020-001010

Figure Lengend Snippet: IL-31 displays potent antitumor activity in vivo. Ten-week-old C57BL/6 female mice were implanted with PyMT cells in the mammary fat pad. After 3 days, mice were intraperitoneally injected with IL-31-L-IgG or control IgG twice weekly (100 µg/inj.). In parallel, another group was implanted with mini-osmotic pumps that delivered purified IL-31 at a dose of 17 µg/day. Control mice were implanted with empty pumps (n=4–5 mice/group). (A) PyMT tumor growth was measured biweekly. (B, C) At endpoint, tumors from mice treated with IL-31-L-IgG or IgG were removed and prepared as single cells. Lymphoid (B) and myeloid (C) populations were analyzed by flow cytometry. (D) Th (CD4 + ) cells, macrophages (CD11b + F4/80 + ), and PMN-MDSCs (Ly6C dim Ly6G + ) were isolated from the spleen of PyMT tumor-bearing C57Bl/6 mice (n=3). RNA was extracted and IL-31Ra mRNA expression was analyzed by RT-qPCR. Relative mRNA levels are present. Statistical significance was assessed by one-way analysis of variance, followed by Tukey post-hoc test for (A) and (D), or by unpaired two-tailed t-test for (B) and (C). Significant p values are shown. CTLs, cytotoxic T lymphocytes; IL, interleukin; IL-31Ra, IL-31 receptor A; MDSCs, myeloid-derived suppressor cells; ns, non-significant; PMN, polymorphonuclear; RT-qPCR, real-time quantitative PCR; TAM, tumor-associated macrophages; Th, T helper.

Article Snippet: The cells were acquired by time of flight mass cytometry (CyTOF) (Fluidigm, South San Francisco, CA, USA) as previously described.

Techniques: Activity Assay, In Vivo, Injection, Control, Purification, Flow Cytometry, Isolation, Expressing, Quantitative RT-PCR, Two Tailed Test, Derivative Assay, Real-time Polymerase Chain Reaction