Dan11mag

Manufactured by Thermo Fisher Scientific

The Dan11mag is a laboratory equipment product designed for magnetic separation. It utilizes a powerful magnetic field to efficiently separate magnetic particles from liquid samples. The core function of the Dan11mag is to facilitate the separation of target analytes or components from complex mixtures, enabling researchers and scientists to extract and purify specific materials for further analysis or processing. No additional details or interpretations about the intended use of this product are provided.

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Lab products found in correlation

Cytofix cytoperm, by BD (2 mentions) Bcl 10c4, by BioLegend (1 mentions) Thr202 tyr204 d13.14.4e, by Cell Signaling Technology (1 mentions) Mp6 xt22, by BD (1 mentions) Foxp3 staining buffer set, by Thermo Fisher Scientific (1 mentions) Golgistop, by BD (1 mentions) Perm wash buffer, by BD (1 mentions) Cytofix cytoperm buffer, by BD (1 mentions) Tim 3, by BD (1 mentions) Lag 3, by BD (1 mentions) Anti cd16 32, by BD (1 mentions) Lsr 2, by FlowJo (1 mentions) Foxp3 transcription factor staining buffer set, by Thermo Fisher Scientific (1 mentions) Phosflow lyse fix buffer, by BD (1 mentions) M89 61, by BD (1 mentions) Phosflow perm buffer 3, by BD (1 mentions) Ebio4b10, by Thermo Fisher Scientific (1 mentions) Lsrfortessa, by BD (1 mentions) Foxp3 staining buffer kit, by Thermo Fisher Scientific (1 mentions) Lsrii flow cytometer, by BD (1 mentions)

5 protocols using dan11mag

Intracellular Cytokine and Protein Staining

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For intracellular cytokine staining (ICS), cells stimulated with indicated stimuli in the presence of GolgiStop (BD Biosciences) were stained for cell-surface markers, fixed and permeabilized using Cytofix/Cytoperm buffer (BD Biosciences) and then stained with fluorochrome-conjugated Abs to IFN-γ (XMG1.2; eBioscience; 1:300) and TNF-α (MP6-XT22; BD Biosciences; 1:300) using Perm/Wash buffer (BD Biosciences). The same ICS protocol was used for analysing Bcl-2 and CD107a expression with fluorochrome-conjugated Abs to Bcl-2 (BCL/10C4; Biolegend; 1:200) and CD107a (1D4B; eBioscience; 1:300). Intracellular staining for T-bet and Eomes expression was performed with Foxp3 Staining Buffer Set (eBioscience) according to the manufacturer's instructions using fluorochrome-conjugated Abs to T-bet (4B10; 1:200) and Eomes (Dan11mag; all from eBioscience; 1:200). For intracellular staining for p-ERK, B6 SP cells treated with indicated stimuli were fixed with 2% paraformaldehyde at RT for 15 min, followed by permeabilization with ice-cold 90% methanol for 20 min on ice. After a washing step, cells were blocked with PBS containing 2% FBS and incubated with fluorochrome-conjugated Ab to p-ERK (Thr202/Tyr204; D13.14.4E; Cell Signaling Technology; 1:100), followed by repeated washes and continued incubation for 15 min on ice with fluorochrome-conjugated Abs to cell-surface markers.

Cho J.H., Kim H.O., Ju Y.J., Kye Y.C., Lee G.W., Lee S.W., Yun C.H., Bottini N., Webster K., Goodnow C.C., Surh C.D., King C, & Sprent J. (2016). CD45-mediated control of TCR tuning in naïve and memory CD8+ T cells. Nature Communications, 7, 13373.

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Multiparametric Characterization of CD8+ T Cells

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Single cell suspensions of splenocytes, obtained from mechanically disrupted and red-blood cell lysed spleens, were Fc-blocked with anti-CD16/32 (BD Biosciences, San Diego, CA clone 2.4G2) and immunophenotyped with the following fluorescently-labeled antibodies to cell surface markers (Biolegend): CD8α (53-6.7), CD44 (1M7), PD-1 (J43), Tim-3 (RMT3-23), Lag-3 (C9B7W), and CD45.1 (A20). MHC class I tetramers specific for D^bGP_33–41, D^bNP_396–404, and D^bGP_276–286 were provided by the National Institutes of Health Tetramer Core Facility (Emory University, Atlanta, GA). For fluorescent staining of intracellular markers, splenocytes, after surface staining, were fixed and permeabilized with either Cytofix/Cytoperm (BD Biosciences) or FoxP3/Transcription Factor Staining Buffer set before staining for Granzyme B (GB12, Life Technologies, Carlsbad, CA) or Eomes (Dan11mag, eBioscience, San Diego, CA). Appropriately conjugated isotype-control antibodies were used in all experiments. Samples were acquired on a BD Biosciences LSR-II and analyzed using FlowJo (Treestar, Ashland, OR).

Hosking M.P., Flynn C.T, & Whitton J.L. (2016). TCR independent suppression of CD8+ T cell cytokine production mediated by IFNγ in vivo. Virology, 498, 69-81.

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Multiparametric Analysis of Immune Cells

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2×10⁶ splenocytes were Fc blocked and immuno-phenotyped with fluorescently-labeled antibodies to the following surface proteins (Biolegend and E Biosciences, Ca): CD8α (53–6.7), Thy1.1 (OX-7), Ly5a (A20), CD44 (1M7), CD69 (H1.2F3), CD25 (PC61), CD62L (MEL-14), and CXCR3 (CXCR3-173). Intracellular marker expression was determined following fixation and permeabilization with either Cytofix/Cytoperm (BD Biosciences) or FoxP3/Transcription Factor Staining Buffer Set (eBioscience) with fluorescently-conjugated antibodies to Granzyme B (GB12, Invitrogen), T-bet (4B10, Biolegend), or Eomes (Dan11mag, eBioscience). Appropriately conjugated isotype control antibodies were used in all experiments. Data were acquired on BD Biosciences LSR-II and analyzed using FlowJo (Treestar, OR). Geometric mean fluorescent intensity (gMFI) was normalized to the average expression following sham infection (0 hours) and expressed as fold induction.

Hosking M.P., Flynn C.T, & Whitton J.L. (2014). Antigen-specific naïve CD8+ T cells produce a single pulse of IFNγ in vivo within hours of infection, but without antiviral effect. Journal of immunology (Baltimore, Md. : 1950), 193(4), 1873-1885.

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Multiparameter Analysis of NK Cell Activation

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For analysis of surface markers CD71, CD98, and CD122, cells were stained in PBS containing 2% (wt/vol) FBS with antibodies from eBioscience or BD. Their expression level was presented as net mean fluorescence intensity (ΔMFI), which was determined by subtracting MFI of isotype control, or as fold relative to unstimulated, set as 1. Intracellular staining was used for NK cell transcription factor and phosphorylated proteins. NK cell transcription factors E4BP4, Eomes, and T-bet were stained with anti–mouse E4BP4 antibody (S2M-E19; eBioscience), anti-Eomes (Dan11mag; eBioscience), and anti–T-bet (eBio4B10; eBioscience) before and after overnight stimulation with IL-15–IL-15R complex according to the manufacturer’s instruction (eBioscience), similar to Foxp3 staining, respectively. For detection of phosphorylated signaling proteins, NK cells were fixed with Phosflow Lyse/Fix buffer, followed by permeabilization with Phosflow Perm buffer III (BD) and staining with antibodies to S6 phosphorylated at Serc235 and Serc236 (D57.2.2E; Cell Signaling Technology), Akt phosphorylated at Serc473 (M89-61; BD), and Thrc308 (J1-223.371; BD). Flow cytometry data were acquired on LSRII or LSR Fortessa (BD) and analyzed using FlowJo software (Tree Star). Net mean fluorescence intensity (ΔMFI) was calculated. Expression levels were presented as fold relative to unstimulated, set as 1.

Yang M., Li D., Chang Z., Yang Z., Tian Z, & Dong Z. (2015). PDK1 orchestrates early NK cell development through induction of E4BP4 expression and maintenance of IL-15 responsiveness. The Journal of Experimental Medicine, 212(2), 253-265.

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Multiparameter Flow Cytometry Analysis

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Splenocytes were stained with a combination of fluorescently labeled monoclonal antibodies (MAb) specific for CD8α (53-6.7), CD8β (YTS156.7.7), Vα2 TCR (B20.1), Ly5.1 (A20), Thy1.1 (HIS51), CD44 (IM7), CD127 (A7R34), CD62L (MEL-14), CD69 (H1.2F3), IFNAR1 (MAR1-5A3), and CD86 (GL1) for 20 min at 4°C. Cells were fixed with BD Cytofix for 5 min at RT and then resuspended in FACS buffer for collection or permeablized for intracellular transcription factor staining. Cells were permeablized for at least 1 hour at 4°C using the Foxp3 staining buffer kit (eBioscience) followed by intracellular staining for IRF4 (3E4), Eomes (Dan11mag), Tbet (eBio4B10), and granzyme B (GB11).
Intracellular cytokine staining was performed as described previously (14 (link)). Cells were stained with a combination of fluorescently labeled MAbs specific for TNF (MP6-XT22), IFNγ (XMG1.2), and granzyme B (GB11, Invitrogen). Stimulating in presence of CD107a (1D4B) and CD107b (ABL-93) identified cells undergoing antigen-induced degranulation. All MAbs were purchased from eBioscience (San Diego, CA), BioLegend (San Diego, CA), or BD Bioscience (San Diego, CA) unless otherwise noted.
All samples, freshly stained or previously fixed, were acquired using a BD Bioscience LSR II flow cytometer with FACS Diva software. Data were analyzed with FlowJo software (Tree Star Inc., Ashland, OR).

Urban S.L., Berg L.J, & Welsh R.M. (2016). Type 1 interferon licenses naïve CD8 T cells to mediate anti-viral cytotoxicity. Virology, 493, 52-59.

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