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Human GRIN1/GRIN2B Stable Cell Line-HEK293

Cat.No.: CSC-RI0179
Cell Line Description: HEK293-HuGRIN1/GRIN2B cell line is a hypotriploid human cell line, which has been co-transfected with a human glutamate receptor, ionotropic, N-methyl D-aspartate 1 (GRIN1) and a human glutamate receptor, ionotropic, N-methyl D-aspartate 2B (GRIN2B) to allow stably express of the human GRIN1 and GRIN2B. It is an example of a cell line transfected using our proprietary CBTGS gene screening and amplification system.
Background: The human GRIN1/GRIN2B genes encode the cationic-selective, n-methyl-d-aspartic acid receptor-channel expressed in the CNS. NMDA receptors are therapeutic targets in treatment of neurological disorders including seizure, pain, and Alzheimer"s disease.
Growth Properties: Adherent
Morphology: Epithelial
Host Cell: HEK293
Cell Line Validation: 1. Gene expression: qPCR experiments determined specific expression of human GRIN1/GRIN2B.2. Protein expression: GRIN1/GRIN2B expression in this cell line has been validated by WB.3. Functional validation: manual patch clamp.
Application: Therapeutic Area: neurodegeneration & stroke, pain & inflammation, psychiatric disorder, seizure-convulsion.
Sub-type: Ligand-gated cation channels
Propagation: Complete growth medium: DMEM/F12/10% FBSAtmosphere: 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.Incubate cultures at 37°C.Subcultivation Ratio: A subcultivation ratio of 1:6 to 1:10 is recommended Medium Renewal: 2 to 3 times per week
Mycoplasma: Mycoplasma Status: Negative (MycoAlert Kit)
Freeze Medium: Complete growth medium 90%; DMSO, 10%
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: GRIN2B glutamate receptor, ionotropic, N-methyl D-aspartate 2B [ Homo sapiens ]
Official Symbol: GRIN2B
Synonyms: GRIN2B; glutamate receptor, ionotropic, N-methyl D-aspartate 2B; NMDAR2B; glutamate [NMDA] receptor subunit epsilon-2; GluN2B; NR3; glutamate receptor subunit epsilon-2; N-methyl-D-aspartate receptor subunit 3; N-methyl D-aspartate receptor subtype 2B; MRD6; NR2B; hNR3; MGC142178; MGC142180;
Gene ID: 2904
mRNA Refseq: NM_000834
Protein Refseq: NP_000825
MIM: 138252
UniProt ID: Q13224
Chromosome Location: 12p12
Pathway: Activation of NMDA receptor upon glutamate binding and postsynaptic events, organism-specific biosystem; Alzheimers disease, organism-specific biosystem; Alzheimers disease, conserved biosystem; Amphetamine addiction, organism-specific biosystem; Amphetamine addiction, conserved biosystem; Amyotrophic lateral sclerosis (ALS), organism-specific biosystem; Amyotrophic lateral sclerosis (ALS), conserved biosystem;
Function: D2 dopamine receptor binding; N-methyl-D-aspartate selective glutamate receptor activity; beta-catenin binding; cell adhesion molecule binding; drug binding; extracellular-glutamate-gated ion channel activity; glycine binding; ion channel activity; ionotropic glutamate receptor binding; metal ion binding; neurotransmitter binding; protein binding; protein heterodimerization activity; receptor activity; receptor binding; transporter activity; zinc ion binding;

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Optional requirements on this protein    +Expand
C-fusion    N-fusion   Non-tagged
His    GST   Fc   Others
<1.0 eu/μg    <0.1 eu/μg   <0.01 eu/μg   Not required
Monomer Isolation    Dimer Isolation    Not required
>80% by SDS-PAGE    >90% by SDS-PAGE   >95% by SDS-PAGE   Others

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