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Human S1PR1-tGFP Stable Cell Line-HepG2

Cat.No.: CSC-RG0027
Cell Line Description: HepG2-HuS1PR1-tGFP cell line is a human liver carcinoma cell line, which has been transfected with a Human Sphingosine-1-phosphate receptor 1(S1P1R) tagged in the C-terminus with tGFP to allow stably express of the human S1P1R tagged in the C-terminus with tGFP. It is an example of a cell line transfected using our proprietary CBTGS gene screening and amplification system.
Background: This gene encodes S1P1 receptor which is a member of the G protein-coupled receptor superfamily (S1P1-5) originally known as EDG receptors. Through the interaction with this receptor family, sphingosine-1-phosphate stimulates diverse cellular responses, including cytoskeletal changes, proliferation and migration. The S1P1R is widely expressed including on endothelial cells. The physiological effects of S1P1R have been described in immune and vascular systems
Growth Properties: Adherent
Morphology: Epithelial
Host Cell: HepG2
Cell Line Validation: 1. Gene expression: RT-PCR experiments determined specific expression of human S1P1R.2. Functional validation: Activation and Internalization assay for S1P1R-tGFP
Sub-type: Lysophospholipid
Propagation: Complete growth medium: The base medium for this cell line is ATCC-formulated Eagle"s Minimum Essential Medium, Catalog No. 30-2003. To make the complete growth medium, add the following components to the base medium: fetal bovine serum to a final concentration of 10%.Atmosphere: air, 95%; carbon dioxide (CO2), 5% Temperature: 37.0°C
Starting Cells From Frozen Cell Stock: 1. Remove the packaging cell lines from liquid nitrogen and carry out a quick thaw. Float the cells in the 37°C water bath for 2 minutes until nearly (80%) thawed. Once cells are thawed, it is important to dilute the cells 1:10 in growth media immediately to reduce the potentially toxic effects of the DMSO preservative on the cells.2. Clean the outside of the vial with 70% ethanol before opening.3. Remove the cells from the vial and add slowly into a 15ml conical tube containing 10 ml pre-warmed media. 4. Centrifuge for 3 minutes 1000 xg to pellet cells and remove the supernatant.5. Add 14 ml of media and transfer cells to a T25 flask or a 100 mm culture dish.6. Place the cells in the 37°C incubator with 5% CO2.7. Allow incubation for 3-4 days to reach confluence. The cells will re-attach to the surface over a period of several days in culture at 37°C.
Subculturing: 1. Remove and discard culture medium.2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin-0.53 mM EDTA solution to remove all traces of serum that contains trypsin inhibitor.3. Add 2.0 to 3.0 ml of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes).Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37°C to facilitate dispersal.4. Add 6.0 to 8.0 ml of complete growth medium and aspirate cells by gently pipetting.5. Add appropriate aliquots of the cell suspension to new culture vessels.6. Incubate cultures at 37°C.Subcultivation Ratio: A subcultivation ratio of 1:4 to 1:6 is recommended Medium Renewal: Twice per week
Mycoplasma: Mycoplasma Status: Negative (MycoAlert Kit)
Freeze Medium: Complete growth medium 90%; DMSO, 5%
Storage: Liquid nitrogen
Preservation: 1. Detach cells from culture dish according to the Sub-Culture Procedure.2. Resuspend cells at a density of 5 x 10^6 cells/mL in freeze medium.Note: A T-75 culture flask typically yields enough cells for preparing two frozen vials.3. Aliquot 1 mL cells into cryogenic vials.4. Place vials in a freezing container and store at –80 °C overnight.5. Transfer vials to liquid nitrogen for long term storage. If properly stored, cells should remain stable for years.
Safety Considerations: The following safety precautions should be observed.1. Use pipette aids to prevent ingestion and keep aerosols down to a minimum.2. No eating, drinking or smoking while handling the stable line.3. Wash hands after handling the stable line and before leaving the lab.4. Decontaminate work surface with disinfectant or 70% ethanol before and after working with stable cells.5. All waste should be considered hazardous.6. Dispose of all liquid waste after each experiment and treat with bleach.
Ship: Dry ice
Gene Name: S1PR1 sphingosine-1-phosphate receptor 1 [ Homo sapiens ]
Official Symbol: S1PR1
Synonyms: S1PR1; sphingosine-1-phosphate receptor 1; EDG1, endothelial differentiation, sphingolipid G protein coupled receptor, 1; sphingosine 1-phosphate receptor 1; CD363; D1S3362; edg 1; S1P receptor 1; S1P receptor Edg-1; sphingosine 1-phosphate receptor EDG1; sphingosine 1-phosphate receptor Edg-1; endothelial differentiation G-protein coupled receptor 1; endothelial differentiation, sphingolipid G-protein-coupled receptor, 1; EDG1; S1P1; ECGF1; EDG-1; CHEDG1; FLJ58121;
Gene ID: 1901
mRNA Refseq: NM_001400
Protein Refseq: NP_001391
MIM: 601974
UniProt ID: P21453
Chromosome Location: 1p21
Pathway: Class A/1 (Rhodopsin-like receptors), organism-specific biosystem; Fc-epsilon receptor I signaling in mast cells, organism-specific biosystem; G alpha (i) signalling events, organism-specific biosystem; GPCR downstream signaling, organism-specific biosystem; GPCR ligand binding, organism-specific biosystem; GPCRs, Other, organism-specific biosystem; Lysosphingolipid and LPA receptors, organism-specific biosystem;
Function: G-protein coupled receptor activity; receptor activity; signal transducer activity; sphingolipid binding; sphingosine-1-phosphate receptor activity;

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