According to the literature, the spleen of the mice was used to prepare single-cell suspension (18 (link)). Then, the effects of SMI on mice T helper (Th1, Th2, Th17) cells and regulatory T (Treg) cells were determined using flow cytometry. In brief, the spleen of mice was digested and single-cell suspensions were stimulated with 50 ng/mL PMA and 1 μg/mL ionomycin in the presence of BD GolgiPlug (Leuko Act Cktl with GolgiPlug, BD Biosciences, San Jose, CA, USA) for 10 hours. Cells were fixed and permeabilized using BD Transcription Factor Buffer Set (BD Biosciences, San Jose, CA, USA) followed by staining with fluorescence-labeled antibodies CD3e-BV605 (145-2C11), CD4-APC-H7 (GK1.5), CD8a-BV510 (53-6.7), CD25-BV421 (PC61), FoxP3-Alexa 647 (MF23), IFN -γ-FITC (XMG1.2), IL-4-PE-Cy7 (11B11) and IL-17A-PE (TC11-18H10). All fluorescence-labeled monoclonal antibodies were purchased from BD Pharmingen. Flow cytometry was performed on a Beckman Coulter CytoFlex flow cytometry system.
Leuko act cktl with golgiplug
Manufactured by BD
Sourced in United States
Leuko Act Cktl with GolgiPlug is a laboratory equipment product. It is used for the detection and analysis of leukocyte activation. The product contains the necessary components to perform this specific function.
Automatically generated - may contain errors
Lab products found in correlation
Transcription factor buffer set, by BD (2 mentions)
Cytoflex flow cytometry system, by Beckman Coulter (1 mentions)
Ifn γ fitc xmg1, by BD (1 mentions)
Golgiplug, by BD (1 mentions)
Mouse th1 th2 th17 cba kit, by BD (1 mentions)
Cytofix cytoperm soln kit, by BD (1 mentions)
Mouse beta actin antibody, by Proteintech (1 mentions)
Fitc anti cd4 antibody, by BD (1 mentions)
Pe anti foxp3 antibody, by Thermo Fisher Scientific (1 mentions)
Myeloperoxidase mpo kit, by Nanjing Jiancheng (1 mentions)
Bca protein determination kit, by Beyotime (1 mentions)
Fixable viability stain 780, by BD (1 mentions)
Trizol reagent, by Takara Bio (1 mentions)
Sybr premix ex taq, by Takara Bio (1 mentions)
Reactive oxygen species assay kit, by Beyotime (1 mentions)
Ros kit, by Nanjing Jiancheng (1 mentions)
Dextran sulfate sodium (dss), by MP Biomedicals (1 mentions)
Glutathione gsh kit, by Nanjing Jiancheng (1 mentions)
D biotin, by Thermo Fisher Scientific (1 mentions)
Pe anti human hla a2 antibody, by BioLegend (1 mentions)
3 protocols using leuko act cktl with golgiplug
1
Evaluation of SMI Effects on Murine T Cells
2
Pharmacological Targeting of Th17/Treg Imbalance in Murine Colitis
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Luteolin and DTT were obtained from Aladdin (China). 1,4-butanediol diacrylate (BDD) was purchased from TCI (Shanghai, China). Dextran sulfate sodium (DSS, 35000 Da) was purchased from MP Biomedical (USA). Mouse Th1/Th2/Th17 CBA Kit, Cytofix/CytopermSoln Kit, Leuko Act Cktl with GolgiPlug, Transcription Factor Buffer Set, Fixable Viability Stain 780, FITC-Anti-CD4 antibody, BV605-Anti-IL-17A antibody, APC-Anti–INF–γ antibody and PE-Anti-IL-4 antibody were purchased from BD Biosciences (San Diego, USA). PE-Anti-FOXP3 antibody was purchased from eBiosciences (San Diego, CA). Myeloperoxidase (MPO) kit, glutathione (GSH) kit and ROS kit were supplied by Nanjing Jiancheng Bioengineering Institute (Nanjing, China). The BCA protein determination kit and Reactive Oxygen Species Assay Kit were purchased from Beyotime Biotechnology (China). Mouse Beta Actin antibody, Mouse Occludin antibody and Mouse ZO-1 antibody for western blotting were purchased from Proteintech (China). SYBR Premix Ex Taq™, Trizol reagent and PrimeScript™ RT 131 Master Mix were obtained from TaKaRa Bio (China). All other reagents are commercially available and can be used directly. All the solvents used were of analytical grade and were procured from Sinopharm (China).
3
Comprehensive Immunologic Assay Protocol
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Antibody were purchased from BioLegend, including FITC anti-human HLA-A2 antibody (Cat#343303), PE anti-human HLA-A2 antibody (Cat#343305), anti-human CD28 antibodies (Cat#302901), APC anti-human-CD69 (Cat#310909), APC anti-human CD8 (Cat#344721), PerCP anti-human IFN-γ (Cat#502524), and FITC anti-human granzyme B (Cat#515403). Flex-T monomer (Cat#280003) and PE conjugated streptavidin (Cat#405203) were purchased from BioLegend. Ficoll-Paque PLUS (Cat#17144003) was from GE and DMSO (Cat#D2650) was from Sigma-Aldrich. IMDM (Cat#SH30228.01) and penicillin-streptomycin (Cat#SV30010) were from HyClone, and FBS (Cat#S711-001S) was purchased from LONSERA. PBS (Cat#C10010500BT) was from Gibco (Waltham, Massachusetts, USA). D-biotin (Cat#2110450) was purchased from Invitrogen (Waltham, Massachusetts, USA), and Mitomycin C (Cat#50-07-7) was from Sinochem Holdings (Beijing, China). CFSE (Cat#T6802) was from Targetmol (Boston, Massachusetts, USA). EasySep Human CD8+ T Cell Isolation Kit (Cat#17953) was from Stem Cell Technologies (Vancouver, British Columbia, Canada), and IL-2 (recombinant human interleukin-2(125Ala) injection) was from SL PHARM (Beijing, China). Leuko act cktl with GolgiPlug (Cat#550583) was purchased from BD.
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