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Human LPAR1 Stable Cell Line-RH7777

Cat.No.: CSC-RG0870
Cell Line Description: RH7777-HuLPAR1 cell line is from Rattus sp. liver hepatoma buffalo, which has been transfected with a human lipid growth factor lysophosphatidic acid (LPAR1) to allow stably express of the human LPAR1. It is an example of a cell line transfected using our proprietary CBTGS gene screening and amplification system.
Background: Lysophosphatidic acid (LPA) is a small, bioactive phospholipid that mediates multiple cellular responses, including proliferation, differentiation, motility, and survival in both normal physiology and disease. The extracellular actions of LPA are mediated primarily through its interaction with five G protein-coupled receptors which include LPA1(EDG2), LPA2(EDG4), LPA3(EDG7), LPA4 (GPR23 or P2Y9) and LPA5 (GPR92). Among them, LPA1 (EDG2) is most widely expressed in normal tissue during growth and development. After LPA stimulation, LPA1 (EDG2) couples to Gi and G12/13 and was reported to play an important role in development of chronic neuroimmune disease and initiation of neuropathic pain.
Growth Properties: Loosely adherent
Morphology: Epithelial
Host Cell: RH7777
Cell Line Validation: 1. Gene expression: qPCR experiments determined specific expression of human LPAR1.2. Protein expression: LPAR1 expression in this cell line has been validated by WB.3. Functional validation: binding, cAMP assay.
Stability: 12 passages of continuous culture (6 weeks) at 2 passages/week
Sub-type: Lysophospholipid
Propagation: Complete growth medium: DMEM, 10% FBS, 0.5 mg/mL G418Atmosphere: 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 culture medium with floating cells to a centrifuge tube.2. If any cells are attached, tap flask gently or if necessary add 2.0 to 3.0 ml of 0.25% Trypsin-0.53 mM EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed. 3. Add 2.0 to 3.0 ml of complete growth medium and aspirate cells by gently pipetting.4. To remove trypsin-EDTA solution, transfer cell suspension to the centrifuge tube with the medium and cells from step #1 and spin at approximately 125 xg for 5 to10 minutes.5. Discard supernatant and resuspend cells in fresh growth medium. Add appropriate aliquots of cell suspension to new culture vessels.6. Place culture vessels in incubators at 37°C. Subcultivation Ratio: A subcultivation ratio of 1:4 to 1:6 weekly is recommended Medium Renewal: Add medium every 2 to 3 days, do not discard floating cells.
Mycoplasma: Mycoplasma Status: Negative (MycoAlert Kit)
Freeze Medium: Complete growth medium 95%; 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: LPAR1 lysophosphatidic acid receptor 1 [ Homo sapiens ]
Official Symbol: LPAR1
Synonyms: LPAR1; lysophosphatidic acid receptor 1; EDG2, endothelial differentiation, lysophosphatidic acid G protein coupled receptor, 2; edg 2; Gpcr26; GPR26; LPA1; Mrec1.3; rec.1.3; vzg 1; LPA-1; LPA receptor 1; ventricular zone gene 1; lysophosphatidic acid receptor Edg-2; endothelial differentiation, lysophosphatidic acid G-protein-coupled receptor, 2; EDG2; VZG1; edg-2; vzg-1;
Gene ID: 1902
mRNA Refseq: NM_001401
Protein Refseq: NP_001392
MIM: 602282
UniProt ID: Q92633
Chromosome Location: 9q
Pathway: Class A/1 (Rhodopsin-like receptors), organism-specific biosystem; G alpha (i) signalling events, organism-specific biosystem; G alpha (q) signalling events, organism-specific biosystem; GPCR downstream signaling, organism-specific biosystem; GPCR ligand binding, organism-specific biosystem; Gap junction, organism-specific biosystem; Gap junction, conserved biosystem;
Function: G-protein alpha-subunit binding; G-protein coupled receptor activity; PDZ domain binding; lysophosphatidic acid receptor activity; phospholipid binding; protein binding; receptor activity; signal transducer activity;

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