Cd4 cd62l isolation kit

Manufactured by Miltenyi Biotec

The CD4+CD62L+ isolation kit is a lab equipment product that allows for the isolation and enrichment of CD4+ T cells expressing the CD62L (L-selectin) surface marker. This kit facilitates the separation of this specific T cell population from a complex sample, such as peripheral blood or tissue, through the use of magnetic beads or columns.

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Lab products found in correlation

Anti ifn γ, by BD (2 mentions) Cd11c microbeads, by Miltenyi Biotec (1 mentions) Anti tgf β, by R&D Systems (1 mentions) Intracellular fixation and permeabilization buffer set, by Thermo Fisher Scientific (1 mentions) Anti il 4, by BD (1 mentions) Interleukin 6 (il 6), by Fujifilm (1 mentions) Live dead fixable aqua dead cell stain, by Thermo Fisher Scientific (1 mentions) Anti cd3, by BD (1 mentions) Anti cd28, by BD (1 mentions) Anti mouse cd8 microbeads, by Miltenyi Biotec (1 mentions) Midimacs magnet, by Miltenyi Biotec (1 mentions) Facsaria 1, by BD (1 mentions) Anti mouse dx5 microbeads, by Miltenyi Biotec (1 mentions) Balb c mice, by Charles River Laboratories (1 mentions) Anti cd3, by BioLegend (1 mentions) Anti cd28, by BioLegend (1 mentions)

Close spelling variants of this product

Mouse CD4+CD62L+ T Cell Isolation Kit II Mouse CD4+ CD62L+ T Cell Isolation Kit CD4+CD62L+ T Cell Isolation Kit II CD4+CD62L+ T cell isolation kit Magnetic CD4+CD62L+ T cell isolation kit MACS CD4+CD62L+ T-cell isolation kit CD62L Isolation kits

6 protocols using cd4 cd62l isolation kit

Isolation of Murine Naive CD4+ and CD8+ T Cells

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For purification of pan-CD4 +and CD8+T cells, splenocytes were cleared of RBCs by hypotonic lysis, and DC populations were depleted with CD11c microbeads (Miltenyi Biotec). T cells were then purified with CD4 or CD8 microbeads.
CD4 +naïve T cells were purified by two strategies. In most experiments, CD4 +cells were pre-enriched from pooled splenocytes and lymph nodes (LNs) by positive selection with CD4 microbeads or depletion with CD19, CD11b, and CD8 microbeads. Naïve CD4 +CD25 CD44-loCD62L-hi cells were then FACS-sorted to >99% purity. In ribosome profiling experiments and for western blot time courses (Figure 1A), naïve CD4 +cells were purified to >95% purity with the CD4 +CD62L + isolation kit (Miltenyi).

Moore M.J., Blachere N.E., Fak J.J., Park C.Y., Sawicka K., Parveen S., Zucker-Scharff I., Moltedo B., Rudensky A.Y, & Darnell R.B. (2018). ZFP36 RNA-binding proteins restrain T cell activation and anti-viral immunity. eLife, 7, e33057.

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Murine Naive CD4+ T Cell Polarization

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Spleen and lymph nodes from mice were harvested, mechanically disrupted, and passed through a 70-μm filter. Naive CD4⁺ T cells from the sample were purified by magnetic bead separation using the CD4⁺CD62L⁺ isolation kit (Miltenyi Biotec). Cells were plated in 96-well plates coated with anti-CD3 antibody (5 μg/mL, BD, clone 145-2C11) at a density of 3 × 10⁶ cells/mL. Cells were plated in Tfh-conditioning media (RPMI complete media with 10% fetal bovine serum), 10 μg/mL anti-IL4 (BD Biosciences, clone 11B11), 10 μg/mL anti-IFNγ (BD Biosciences, clone XMG1.2), 10 μg/mL anti-TGFβ (R&D Systems, clone 1D11), 30 ng/mL IL6 (Shenandoah Biotechnology), and 50 ng/mL IL21 (Shenandoah Biotechnology) in the presence of soluble anti-CD28 (2 μg/mL, BD, clone 37.51). Polarized cells were harvested for Tfh phenotyping by flow cytometric analysis 4 days after plating.
For surface phenotype and transcription factor analysis, harvested cells were stained with Live/Dead Fixable Aqua dead cell stain (Thermo Scientific Fisher) to exclude dead cells. Cells were subsequently stained for surface markers (CD4, TCRβ, PD1, CXCR5) and then fixed and permeabilized using the Intracellular Fixation and Permeabilization Buffer Set (eBioscience). Fixed cells were stained with antibody to BLC6 (clone 7D1) or isotype control.

Battaglia M., Sunshine A.C., Luo W., Jin R., Stith A., Lindemann M., Miller L.S., Sinha S., Wohlfert E, & Garrett-Sinha L.A. (2023). Ets1 and IL17RA cooperate to regulate autoimmune responses and skin immunity to Staphylococcus aureus. Frontiers in Immunology, 14, 1208200.

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Murine T Cell Differentiation Assay

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Naïve T cells were isolated from the spleen and lymph nodes of mice with a Miltenyi CD4+CD62L+ isolation kit and were cultured for 4 days on plate-bound anti-CD3 (3μg/mL) and anti-CD28 (3μg/mL) ; BD Pharmingen Cats. 553057 and 553294. Th0: anti-IL-4 (10μg/mL); BD Pharmingen Cat. 554432, anti-IFN-γ (10μg/mL) ; BD Pharmingen Cat. 554408; Th1: anti-IL-4 (10μg/mL), IL-12 (10ng/mL); Th17: anti-IL4 (10μg/mL), anti-IFN-γ (10μg/mL), IL-6 (20ng/mL), TGF-β (5ng/mL); Th2: anti-IFN-γ (10μg/mL), IL-4 (10ng/mL), IL-2 (10ng/mL); Th25: anti-IFN-γ (10μg/mL), IL-25 (100ng/mL). For intracellular staining, cells were stimulated with PMA and ionomycin for 5 hours, GolgiStop was added during the last 2 hours of stimulation.

Wu L., Zepp J.A., Qian W., Martin B.N., Yin W., Bunting K.D., Aronica M., Erzurum S, & Li X. (2015). A novel IL-25-signaling pathway through STAT5. Journal of immunology (Baltimore, Md. : 1950), 194(9), 4528-4534.

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Isolation of Lymphocyte Subsets from Mouse Spleen

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Different lymphocyte subsets were purified from splenic mononuclear cells isolated from Balb/c mice (Charles River Laboratories, Wilmington, MA, USA). If necessary, further isolation was performed by magnetic activated cell sorting (MACS; Miltenyi Biotec, Bergisch Gladbach, Germany) or by FACS (FACSAria I, BD Biosciences, San Jose, USA).
Briefly, cell suspension of the spleen was prepared by cutting small pieces and gently pressing through a 100 μm wire mesh. DX5⁺ cells were purified using anti-mouse-DX5⁺ MicroBeads (Miltenyi Biotec). Cells were passed through a MACS column (type LS) attached to a MidiMACS magnet (Miltenyi Biotec). DX5⁺ cells were collected in the positive fraction. DX5⁺ splenocytes were labeled with FITC-conjugated anti-mouse CD3 molecular complex (clone: 17A2, rat IgG2b) and PE-conjugated anti-mouse CD49b (clone: DX5, rat IgM) (all from BD Biosciences) for further DX5⁺NKT cell isolation by FACS sorting. CD4⁺CD62L^high and CD4⁺CD62L_low cells were purified using the CD4⁺CD62L⁺ Isolation Kit (Miltenyi Biotec) and CD8⁺ cells by using anti-mouse-CD8⁺ MicroBeads (Miltenyi Biotec).

Werner J.M., Damian M., Farkas S.A., Schlitt H.J., Geissler E.K, & Hornung M. (2018). Murine DX5+NKT Cells Display Their Cytotoxic and Proapoptotic Potentials against Colitis-Inducing CD4+CD62Lhigh T Cells through Fas Ligand. Journal of Immunology Research, 2018, 8175810.

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Isolation and Differentiation of CD4+ T Cells

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Lung and spleen CD4⁺ T cells were isolated by magnetic-activated cell sorting (MACS). Naive CD4⁺ T cells were isolated using the CD4⁺CD62L⁺ isolation kit (Miltenyi Biotec; ~95% purity) or by fluorescence-activated cell sorting (FACS; ~99% purity). Human CD4⁺ T cells were isolated by MACS from peripheral blood. Cells were cultured in the presence of anti-CD3 (BioLegend 317302; 3 μg/mL) and anti-CD28 (BioLegend 302902; 1 μg/mL) and differentiated to Th1 (with IL-12, 20 ng/mL), Th2 (IL-4, 20 ng/mL), Th9 (IL-4, 20 ng/mL, and TGF-β, 2 ng/mL), Th17 (TGF-β, 10 ng/mL, and IL-6, 30 ng/mL), and Tregs (TGF-β, 10 ng/mL). In some experiments, cells were treated with cTXA₂ (7.5–300 nM), TXB₂ (500 nM), or TP receptor agonist U-46619 (300 nM) or corresponding vehicle.

Li H., Bradbury J.A., Edin M.L., Gruzdev A., Li H., Graves J.P., DeGraff L.M., Lih F.B., Feng C., Wolf E.R., Bortner C.D., London S.J., Sparks M.A., Coffman T.M, & Zeldin D.C. (2024). TXA2 attenuates allergic lung inflammation through regulation of Th2, Th9, and Treg differentiation. The Journal of Clinical Investigation, 134(9), e165689.

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Zfp36 KO CD4+ T Cell Activation

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Naïve CD4 +T cells were purified with the CD4 +CD62L + isolation kit (Miltenyi) and activated in the presence of formalin-fixed DCs, as described above. After 4 hr, 100 μg/ml cycloheximide was added to cultures and briefly incubated at 37°C. Cells were harvested on ice, washed in 1X PBS containing cycloheximide, pelleted, and snap frozen in liquid nitrogen. Four pairs of biological replicates (WT and Zfp36 KO) were analyzed.

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