Anti cd103

Manufactured by Thermo Fisher Scientific

Sourced in United States

Anti-CD103 is a monoclonal antibody that binds to the CD103 (integrin αE) antigen. CD103 is expressed on intraepithelial T cells and a subset of dendritic cells. The core function of Anti-CD103 is to detect and identify cells expressing the CD103 antigen.

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4 protocols using anti cd103

Analyzing Lung-Resident Memory CD8+ T Cells

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Lung-resident memory CD8⁺ T cells were analyzed as previously described (34 (link)). Mice were intranasally infected with H1N1 or H5N1 (2:6) virus. On day 30 postinfection, mice were intravenously injected with 1 μg anti-CD8β antibody 5 minutes before tissue harvest. Lung tissues were perfused with PBS and single-cell suspensions were prepared after digestion with collagenase as described before. Cells were blocked first with FcRγIII/II antibody and stained with H-2D^b restricted tetramer conjugated to fluorophore R-phycoerythrin (PE) (NP_366-374 ASNENMETM). Tetramer-labeled cells were washed and stained with anti-CD4 (2 μg/ml; clone RM4-4; BioLegend), anti-CD3 (2 μg/ml; clone 145-2C11; eBiosciences), anti-CD8a (1 μg/ml; clone 53-6.7; eBiosciences), anti-CD44 (clone IM7; BioLegend), anti-CD103 (2 μg/ml; clone 2E7; eBiosciences), and anti-CD69 (clone H1.2F3; BioLegend). Dead cells were stained with the Live/Dead fixable near-IR staining kit (Life Technologies) in PBS for 15 minutes on ice. Surface-stained samples were fixed with FACS buffer containing 0.1% formaldehyde and analyzed using the BD LSR-II flow cytometer. Data analysis was performed using FlowJo software (Treestar Corp.).

Kandasamy M., Furlong K., Perez J.T., Manicassamy S, & Manicassamy B. (2020). Suppression of Cytotoxic T Cell Functions and Decreased Levels of Tissue-Resident Memory T Cells during H5N1 Infection. Journal of Virology, 94(9), e00057-20.

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Lung and Tumor Immune Cell Isolation

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Lymphocytes were isolated from lung and tumor tissue by digestion with collagenase A (1 mg/ml; Roche) and DNase I (0.5 µg/ml; Roche) in isolation buffer (RPMI 1640 supplemented with 5% FBS, 1% l-glutamine, 1% penicillin-streptomycin, and 10 mM Hepes) for 30 min at 37°C. Cells were filtered through 100-µm cell strainers, washed in isolation buffer, and stained in PBS supplemented with 0.25% BSA, 2 mM EDTA, and 0.1% sodium azide. Antibodies used included anti-CD45, anti-Foxp3, anti–IL-18Rα, anti-ST2, anti-CD62L, anti-CD103, anti–PD-1, anti-GITR, anti–CTLA-4, anti-KLRG1, anti-Ki67, anti-CD5, anti-NK1.1, anti-CD45R (B220), anti–IL-17, anti–IL-4, anti-IFNγ, and anti–Ly-6C (eBioscience); anti-TCRβ, anti-CD3, anti-CD4, anti-CD8, anti-CD127, anti-CD11B, anti-MHCII, and anti-Gr1 (BioLegend); anti-CD44 and anti-CD25 (Tonbo); anti–IL-5 and anti-TNFα (BD PharMingen); and anti-AREG (R&D Systems). For exclusion of dead cells, samples were first stained with Ghost Dye (Tonbo) cell viability reagent. Intracellular staining for cytokines, Foxp3, AREG, Ki-67, and CTLA-4 was performed using the Foxp3/transcription factor staining buffer set (eBioscience) as per manufacturer’s protocol. The H2-K^b OVA_257–264 tetramer was obtained from the NIH Tetramer Core Facility.

Green J.A., Arpaia N., Schizas M., Dobrin A, & Rudensky A.Y. (2017). A nonimmune function of T cells in promoting lung tumor progression. The Journal of Experimental Medicine, 214(12), 3565-3575.

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Multiparametric Flow Cytometry of Immune Cells

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Flow cytometric analyses of leukocytes in the mesenteric lymph nodes (MNL), spleen, kidney, and intestine were performed. The cells were stained with fluorochrome-labeled monoclonal antibodies (anti-CD4, anti-CD25, Fixable Viability Dye, anti-Foxp3, anti-CD45, anti-CD11c, anti-CD103, anti-CX₃CR₁, and anti-Ly6C; all eBioscience, BioLegend, or BD Biosciences [Franklin Lakes, NJ, US]) and analyzed using a four-color flow cytometer (FACSCanto II; BD Biosciences) and FlowJo software (Tree Star Inc., Ashland, CA, USA).

Yang J., Ji G.E., Park M.S., Seong Y.J., Go Y.S., Lee H.Y., Fang Y., Kim M.G., Oh S.W., Cho W.Y, & Jo S.K. (2021). Probiotics partially attenuate the severity of acute kidney injury through an immunomodulatory effect. Kidney Research and Clinical Practice, 40(4), 620-633.

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Flow Cytometric Analysis of T cell Subsets

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Fluorescent-dye-conjugated antibodies were purchased from BD-Pharmingen (anti-CD4, 550954; anti-CD25, 553866; anti-CD103, 557495; anti-IL-17a, 559502; anti-T-bet, 561312) or eBioscience (anti-CD8α, 56-0081; anti-CD44, 56-0441; anti-CD45.1, 25-0453; anti-CD45.2, 47-0454; anti-CD62L, 48-0621; anti-TCR-β, 47-5961; anti-IFN-γ, 25-7311; anti-IL-22, 24-7221; anti-RORγt, 12-6981; anti-Foxp3, 17-5773). Flow cytometry data was acquired on a LSR-II flow cytometer (Becton Dickinson) and analyzed using FlowJo software package (Tree Star). Intracellular staining of Foxp3 was conducted using Foxp3 Mouse Regulatory T cell Staining Kit (eBioscience).
For flow cytometric analysis of cytokine-secreting cells, cells were incubated in the presence of 100ng/ml PMA (Sigma), 500ng/ml Ionomycin (Sigma) for 4.5h and 10μg/ml brefeldin A (BFA) (Sigma) for the last 2.5h prior staning. Cell populations were first stained with antibodies against the indicated cell surface markers, followed by permeabilization in Fix/Perm buffer, and intracellular staining in Perm/Wash buffer (BD Pharmingen).

Reis B.S., Lee K., Fanok M.H., Mascaraque C., Amoury M., Cohn L., Rogoz A., Dallner O.S., Moraes-Vieira P.M., Domingos A.I, & Mucida D. (2015). Leptin receptor signaling in T cells is required for Th17 differentiation. Journal of immunology (Baltimore, Md. : 1950), 194(11), 5253-5260.

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