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Human HRH4-FLAG Stable Cell Line-HEK293T

Cat.No.: CSC-RG0444
Cell Line Description: HEK293T-HuHRH4-FLAG cell line is a hypotriploid human cell line, which has been transfected with a human histamine receptor H4 (HRH4) tagged in the N-terminus with FLAG to allow stably express of the human HRH4 tagged in the N-terminus with FLAG. It is an example of a cell line transfected using our proprietary CBTGS gene screening and amplification system.
Background: Histamine is one of the most studied biomolecules in medicine and is most notably known for its effects on smooth muscle contraction, vascular permeability and regulation of stomach acid. The histamine receptor H4 has been shown to have a role in chemotaxis and mediator release in a variety of immune cells, such as mast cells, eosinophils, dendritic cells, and T cells. The development of potent H4 receptor antagonists has great potential to open up the pathway for new therapeutic treatments in chronic inflammatory diseases, such as bronchial asthma, allergic gastrointestinal disease, and atopic dermatitis.
Growth Properties: Adherent
Morphology: Epithelial
Host Cell: HEK293T
Cell Line Validation: 1. Gene expression: RT-PCR experiments determined specific expression of human HRH4.2. Protein expression: HRH4 expression in this cell line has been validated by WB.3. Functional validation: Functional assays.
Sub-type: Histamine
Propagation: Complete growth medium: DMEM, 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 1:4 to 1:6 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: HRH4 histamine receptor H4 [ Homo sapiens ]
Official Symbol: HRH4
Synonyms: HRH4; histamine receptor H4; histamine H4 receptor; AXOR35; GPCR105; GPRv53; H4R; HH4R; SP9144; pfi-013; G-protein coupled receptor 105; H4; BG26; MGC133027;
Gene ID: 59340
mRNA Refseq: NM_001143828
Protein Refseq: NP_001137300
MIM: 606792
UniProt ID: Q9H3N8
Chromosome Location: 18q11.2
Pathway: Amine ligand-binding receptors, organism-specific biosystem; Class A/1 (Rhodopsin-like receptors), 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; Histamine receptors, organism-specific biosystem;
Function: G-protein coupled receptor activity; histamine receptor activity; receptor activity; signal transducer activity;

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