dLNs (2 inguinal, 2 axillary, 2 brachial, 2 popliteal, and 4 cervical LNs per mouse) were isolated and digested as described (45 (link)) in RPMI 1640 with 2% FBS, 0.8 mg/ml Dispase (Roche Diagnostics), 0.2 mg/ml Collagenase P (Roche Diagnostics), and 0.1 mg/ml DNase I (Sigma-Aldrich). Single cell suspensions were stained in the presence of Fc-block with fluorochrome-conjugated antibodies (all purchased from BioLegend unless otherwise indicated) against the following targets: CD4 (RM4-4), CD45 (30-F11), CD3 (1452C11), TCRb (H57-597), CD44 (IM7), CD62L (MEL-14), CD69 (H1.2F3), gp38 (8.1.1), CD31 (MEK13.3), S1PR1 (713412, R&D Systems). Staining with S1PR1 antibody was performed in the presence of fatty acid–free BSA from Sigma-Aldrich; blood cells were used as S1PR negative controls (17 (link), 46 (link)). Stained cells were analyzed with the FACS Canto II (BD Biosciences) and Flow-Jo software. For ki67 (clone 16A8, BioLegend) intracellular staining, cells were fixed and permeabilized with the Foxp3 Fix/Perm Buffer Set (BioLegend). LN stromal cells were sorted as described (45 (link)) using a BD FACSAria Fusion cell sorter; CD45+ cells were depleted prior to sorting with anti-biotin magnetic beads (Miltenyi Biotec).
S1pr1
Manufactured by Merck Group
Sourced in United States
S1PR1 is a laboratory equipment product manufactured by Merck Group. It functions as a receptor that binds to the signaling molecule sphingosine-1-phosphate, which plays a role in various cellular processes.
Automatically generated - may contain errors
Lab products found in correlation
S1pr3, by Merck Group (2 mentions)
Facscanto 2, by BD (1 mentions)
Dnase 1, by Merck Group (1 mentions)
Anti biotin magnetic beads, by Miltenyi Biotec (1 mentions)
Facsaria fusion cell sorter, by BD (1 mentions)
Dispase, by Roche (1 mentions)
Collagenase p, by Roche (1 mentions)
Foxp3 fix perm buffer set, by BioLegend (1 mentions)
S1pr1, by R&D Systems (1 mentions)
Fluorochrome conjugated antibodies, by BioLegend (1 mentions)
C fos, by Merck Group (1 mentions)
C jun, by Merck Group (1 mentions)
Pdgfr, by Merck Group (1 mentions)
C src, by Merck Group (1 mentions)
Metafectene transfection reagent, by Biontex (1 mentions)
3 protocols using s1pr1
1
Isolation and Analysis of Lymph Node Cells
2
Targeted Gene Silencing by siRNA
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Human siRNAs of scrambled, S1PR1, S1PR3, c-Src, EGFR, PDGFR, p38, p42, JNK1, c-Jun, and c-Fos were from Sigma (St. Louis, MO). Transient transfection of siRNAs was carried out using Metafectene transfection reagent (Biontex, Germany). siRNA (100 nM) was formulated with Metafectene transfection reagent according to the manufacturer’s instruction.
3
siRNA Transfection in HCF Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The SMARTpool-siRNAs of scramble, S1PR1, S1PR3, Gq, Gi, PKCα, p42, p38α, JNK1, and p65 were from Sigma (St. Louis, MO, United States). HCFs cells were plated at 3 cells/ml × 105 cells/ml (1 ml per well) in 12-well culture plates for 24 h, reaching about 80% confluence. Cells were incubated with 0.4 ml of serum-free DMEM/F-12 medium each well after washed once with PBS and once with serum-free DMEM/F-12. The siRNA was prepared and transiently transfected using Lipofectamine 2000 transfection reagent according to the instructions of the manufacturer.
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!