Fluorochrome-conjugated antibodies to CD4 (GK1.5), CD11b (M1/70), CD11c (N418), CD45.1(A20), CD45.2 (104), CD90.1 (OX-7), CD103 (2E7), I-A/I-E (M5/114.15.2), Dectin-1 (RH1), IFN-γ (XMG1.2), IL-10 (JES5-16E3) and IL-17A (TC11-18H10.1) were purchased from BioLegend (San Diego, CA). Anti-mouse Langerin (L31), IL-17F (18F10), IL-22 (1H8PWSR), Foxp3 (FJK-16S) were acquired from eBioscience (San Diego, CA).
Il 17a tc11 18h10.1
Manufactured by BioLegend
Sourced in United States
IL-17A (TC11-18H10.1) is an antibody product from BioLegend. It is a purified anti-mouse interleukin-17A (IL-17A) monoclonal antibody. IL-17A is a pro-inflammatory cytokine produced by various cell types, including Th17 cells.
Automatically generated - may contain errors
Lab products found in correlation
Facscalibur, by BD (4 mentions)
Ifn γ xmg1.2, by BioLegend (4 mentions)
Cd62l mel 14, by BioLegend (3 mentions)
Facscanto, by BD (3 mentions)
Cd11b m1 70, by BioLegend (3 mentions)
Anti tgf β mab 1d11, by R&D Systems (2 mentions)
Ifn γ xmg1.2, by BD (2 mentions)
Ionomycin, by Merck Group (2 mentions)
Ki67 sola15, by Thermo Fisher Scientific (2 mentions)
Rorγt afkjs 9, by Thermo Fisher Scientific (2 mentions)
Il 23r 753317, by R&D Systems (2 mentions)
Gr 1 rb6 8c5, by BioLegend (2 mentions)
Cytofix cytoperm, by BD (2 mentions)
Il 10 jes5 16e3, by BioLegend (1 mentions)
Il 21r fc chimera, by R&D Systems (1 mentions)
Human tgf β, by R&D Systems (1 mentions)
Il 23, by R&D Systems (1 mentions)
Permeabilization buffer, by Thermo Fisher Scientific (1 mentions)
Lsrfortessa, by BD (1 mentions)
Ic fixation buffer, by Thermo Fisher Scientific (1 mentions)
16 protocols using il 17a tc11 18h10.1
1
Multiparameter flow cytometry analysis
2
Murine Immune Cell Staining
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Antibodies to CD3ε (145-2C11), CD28 (37.51), CD11c (HL3), CD44 (IM7), CD62L (MEL-14), human NGFR (C40-1457), IL-4 (11B11), IL-4 (BDV4-1D11), and IFN-γ (XMG1.2) were purchased from BD. Antibodies to CD8α (53–6.7), B220 (RA3-6B2), Thy1.1 (OX-7), NKp46 (29A1.4), and IL-17A (TC11-18H10.1) were purchased from BioLegend. Antibodies to Foxp3 (FJK-16s), IL-17F (eBio18F10), and γδ TCR (UC7-13D5) were purchased from eBioscience. Murine IL-1β, IL-4, and IL-6 were purchased from PeproTech. Human TGF-β, murine IL-21, IL-23, anti–TGF-β mAb (1D11), and IL-21R-Fc chimera were purchased from R&D Systems.
3
Mouse CD1d Tetramer Staining
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Mouse CD1d tetramers coupled to APC and loaded with PBS-57 (an iNKT cell ligand analog to α-GalCer) were provided by the NIH Tetramer Core Facility. Fluorochrome-labeled monoclonal antibodies (mAbs) specific to mouse CD45.1 (A20), CD45.2 (104), TCRβ (H57-597), CD4 (RM4-5), NK1.1 (PK136), Gr-1 (RB6-8C5), CD11b (M1/70), CD69 (H1.2F3), IFN-γ (XMG1.2), IL-4 (11B11), and IL-17A (TC11-18H10.1) were all purchased from BioLegend. c-Maf mAb (sym0F1) was purchased from eBioscience. For intracellular cytokine staining, cells were incubated with PMA (50 ng/ml), ionomycin (500 ng/ml), and brefeldin A (10 µg/ml; Sigma) for 3 h. After preincubation with unconjugated anti-CD16/32 (2.4G2; BioLegend) mAb, cells were stained for surface antigen and then fixed and permeabilized using the IC fixation buffer and permeabilization buffer (eBioscience) followed by incubation with anti-cytokine or isotype control Abs. FACS data were acquired by BD LSRFortessa (BD Biosciences) and analyzed with FlowJo software. For cell sorting, murine naive CD4+CD62Lhi T cells and CD1d/PBS-57+TCRβ+ iNKT cells were sorted by FACSAria IIIu (BD Biosciences).
4
Multiparameter Flow Cytometry Analysis of γδ T Cells
5
Characterization of CD4+ T Cells and Astrocytes
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After 48 h or 96 h of incubation, non-adherent CD4 T cells were removed by washing the co-culture 3 times with PBS (2% FBS). The remaining adherent cells (T cells and astrocytes) were incubated with accutase (Millipore) at 37°C for 40 min, collected and analyzed by flow cytometry. CD4+ cells were stained with the following mouse specific monoclonal antibodies: CD3 (clone 17A2), CD4 (GK1.5), MHCII (M5/114.15.2), CD39 (24DMS1), Rorg (gamma, greek letter) t (lower case) (AFKJS-9), IFN-γ (XMG1.2), CD25 (PC61 5), FoxP3 (FJK-165), CD44 (IM7), CD62L (MEL-14) (eBioscience), CD73 (TY11.8) and IL-17A (TC11-18H10.1) (Biolegend). Astrocytes were stained for CD11b (M1/70) (Biolegend), VCAM-1 (429) (BD Bioscience), ICAM-1 (YN1/1.7.4) (eBioscience). All samples were acquired with a FACSCalibur or FACSCanto (Becton Dickinson) and analyzed with FlowJo software (FlowJo v10.0.7). IL-6 protein concentration was determined using the DuoSet ELISA Development System according to the manufacturer's instructions (R&D Systems).
6
Multiparameter Flow Cytometry Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Whole spinal cord infiltrated cells or 2.0×105 lymphocytes were stained with 2
µg/ml monoclonal antibodies (mAbs) after Fcγ receptor blocking with
2.4G2 Ab. After cell surface staining, cells were fixed with 4% paraformaldehyde and
permeabilized with 0.1% saponin, and then intracellular staining was performed. Anti-mouse
CD11c (N418 or HL3), CD4 (RM4-5 or GK1.5), CD8 (53-6.7), CD40 (3/2, 3), CD80 (16-10A1),
CD86 (GL-1), major histocompatibility complex class II (MHC class II) (M5/114.15.2),
granulocyte-differentiation antigen-1 (Gr-1) (RB6-8C5), IFN-γ (XMG1.2) and IL-17A
(TC11-18H10.1) were purchased from Biolegend (USA). Phcoerythrin (PE)-conjugated IL-17A
(TC11-18H10) and biotin-conjugated anti-mouse OX40 ligand (OX40L) (RM134L) were purchased
from BD Pharmingen (USA). PE/Cy7-conjugated streptavidin was purchased from BD Bioscience
(USA). Stained cells were detected with a FACSCanto II Flow Cytometer (Becton, Dickinson
and Company, USA) and analyzed with BD FACS Diva (Becton, Dickinson and Company) and
FlowJo software (Tree Star, USA). Dead cells which stained by 7-Amino Actinomycin D
(7-AAD) were excluded.
µg/ml monoclonal antibodies (mAbs) after Fcγ receptor blocking with
2.4G2 Ab. After cell surface staining, cells were fixed with 4% paraformaldehyde and
permeabilized with 0.1% saponin, and then intracellular staining was performed. Anti-mouse
CD11c (N418 or HL3), CD4 (RM4-5 or GK1.5), CD8 (53-6.7), CD40 (3/2, 3), CD80 (16-10A1),
CD86 (GL-1), major histocompatibility complex class II (MHC class II) (M5/114.15.2),
granulocyte-differentiation antigen-1 (Gr-1) (RB6-8C5), IFN-γ (XMG1.2) and IL-17A
(TC11-18H10.1) were purchased from Biolegend (USA). Phcoerythrin (PE)-conjugated IL-17A
(TC11-18H10) and biotin-conjugated anti-mouse OX40 ligand (OX40L) (RM134L) were purchased
from BD Pharmingen (USA). PE/Cy7-conjugated streptavidin was purchased from BD Bioscience
(USA). Stained cells were detected with a FACSCanto II Flow Cytometer (Becton, Dickinson
and Company, USA) and analyzed with BD FACS Diva (Becton, Dickinson and Company) and
FlowJo software (Tree Star, USA). Dead cells which stained by 7-Amino Actinomycin D
(7-AAD) were excluded.
7
Mesenteric Lymph Node Cell Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Mesenteric lymph nodes (MLNs) were manually dissected, and single-cell suspensions were made by passing cells through a 70μM cell strainer in RPMI 1640 media (Life Technologies) containing 10% FCS (Hyclone), 2 mM l -glutamine (Life Technologies), and 100 U/ml penicillin and 100 μg/ml streptomycin (Life Technologies).
Cells were stained directly ex vivo for Foxp3 measurements or, for intracellular cytokine measurements, restimulated with 0.5 μg/ml PMA and 1 μg/ml ionomycin for 3.5 h, with 10 μg/ml brefeldin A included for the final 2.5 h. Cells were stained for 20 min at 4°C with Abs to the surface marker CD4 (RM4-5; BD Pharmingen), and CD103 conjugated to biotin (M290; BD Pharmingen) followed by PerCP-streptavidin (BD Pharmingen). Cells were fixed according to the manufacturer’s instructions with Fix/Perm (eBiosciences) for Foxp3 measurements, or Cytofix/Cytoperm (BD) for intracellular cytokine measurements, and then stained for 20 min at 4°C with Abs to Foxp3 (FJK-16s; eBioscience), IL-4 (11B11; BioLegend), IFN-γ (XMG1.2; BioLegend), IL-17A (TC11-18H10.1; BioLegend), or the relevant isotype controls. Marker expression was measured on FACSCanto (BD) or LSR II (BD) flow cytometers, and data were analyzed using FlowJo software (TreeStar).
Cells were stained directly ex vivo for Foxp3 measurements or, for intracellular cytokine measurements, restimulated with 0.5 μg/ml PMA and 1 μg/ml ionomycin for 3.5 h, with 10 μg/ml brefeldin A included for the final 2.5 h. Cells were stained for 20 min at 4°C with Abs to the surface marker CD4 (RM4-5; BD Pharmingen), and CD103 conjugated to biotin (M290; BD Pharmingen) followed by PerCP-streptavidin (BD Pharmingen). Cells were fixed according to the manufacturer’s instructions with Fix/Perm (eBiosciences) for Foxp3 measurements, or Cytofix/Cytoperm (BD) for intracellular cytokine measurements, and then stained for 20 min at 4°C with Abs to Foxp3 (FJK-16s; eBioscience), IL-4 (11B11; BioLegend), IFN-γ (XMG1.2; BioLegend), IL-17A (TC11-18H10.1; BioLegend), or the relevant isotype controls. Marker expression was measured on FACSCanto (BD) or LSR II (BD) flow cytometers, and data were analyzed using FlowJo software (TreeStar).
8
Multicolor Flow Cytometry of Immune Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Membrane and intracellular staining of MLN or epithelial cells were performed as described.22 The following antibodies were used: CD4 (GK1.5, BioLegend), CD8α (53–6.7, BD Biosciences), CD16/CD32 (93, BioLegend), IL-17A (TC11-18H10.1, BioLegend), RORγt (Q31-378, BD Biosciences), IFNγ (XMG1.2, BD Biosciences), IL-10RA (1B1.3a, BioLegend), and Ep-CAM (G8.8, BioLegend).
9
Intracellular Cytokine Staining and Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For intracellular cytokine staining, cells were restimulated for 4 h with 50 ng/ml phorbol 12-myristate 13-acetate and 1 µM ionomycin (Sigma-Aldrich) in the presence of GolgiStop (BD). Cells were fixed with Cytofix/Cytoperm Fixation and Permeabilization Solution (BD) and stained for intracellular cytokines. Cells were analyzed on a FACSCalibur (BD). Antibodies to the following antigens were used: CD201 (EPCR; eBio1560; eBioscience), CD4 (RM4-5; BioLegend), IL-17A (TC11-18H10.1; BioLegend), IFN-γ (XMG1.2; BioLegend), CD62L (MEL-14; BioLegend), CD44 (IM7; BioLegend), IL-1R (JAMA-147; BioLegend), and GM-CSF (MP1-22E9; BioLegend). All flow cytometric data were analyzed using FlowJo Software (Tree Star). IL-17A concentration in the supernatant was determined by ELISA using standard protocols.
10
Flow Cytometric Analysis of Skin-Infiltrating T Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Fluorochrome-labeled mAbs including anti-mouse CD3 (17A2, RRID: AB_1595492), γδTCR (GL3, RRID: AB_1595492), Vγ4 (UC3-10A6, RRID: AB_10569353), Vδ4 (GL2, RRID: AB_1877234), CD45 (30-F11, RRID: AB_312973), CD11b (M1/70, RRID: AB_312791), Gr-1 (RB6-8C5, RRID: AB_313377), and IL-17A (TC11-18H10.1, RRID: AB_2125010) and IL1R1 (DIH9, RRID: AB_2687367) were purchased from Biolegend. Anti-human γδTCR (REA591, Cat # 130-113-508) was purchased from Miltenyi Biotec and anti-human CD3 (UCHT1, Cat # 550795) was purchased from BD bioscience. For intracellular IL-17 staining, mouse skin cells or LN cells were treated with GolgiPlug (Biolegend) at 37°C for 4 h. Cells were first blocked with anti-CD16/32 and then stained with different cell surface Abs at 4°C for 20 min. Cells were then fixed, permeabilized and stained intracellularly at 4°C overnight for IL-17A. The relevant isotype control mAbs were also used. Samples were harvested with BD FACS Canto and analyzed with FlowJo software (TreeStar).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!