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Immortalized Mouse Cranial Suture Cells (iMSu)

SKU : BHC10900735

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Applied Biological Materials (abm) Inc.

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Mfr.No.	T0227
Description	Immortalized Mouse Cranial Suture Cells (iMSu) are immortalized through infection with non-replicating retrovirus carrying the loxP-flanked SV40 large T antigen. In vitro, the cells can be genetically manipulated for further research. Thus it is a reliable tool for research in mouse cranial suture biology and molecular signalling mechanism implicated in cranial sutures pathophysiology such as craniosynostosis.
Cell Type	Immortalized Cells
Biosafety Level	2
Depositor	University of Chicago
Disclaimer	1. All test parameters provided in the CoA are conducted using abm's standardized culture system and The stated values may vary under the end-user's culture conditions. Please verify that the product is suitable for your studies by referencing published papers or ordering RNA (0.5 ?g, Cat.# C207, $450.00) or cell lysate (100 ?g, Cat.# C206, $600.00) to perform preliminary experiments, or alternatively use our Gene Expression Assay Service (Cat# C138). All sales are final. 2. We recommend live cell shipments for ease of cell transfer and this option can be requested at the time of order placement. Please note that the end-user will need to evaluate the feasibility of live cell shipment by taking into account the final destination's temperature variation and its geographical location. 3. All of abm's cell biology products are for research use ONLY and NOT for therapeutic/diagnostic applications. abm is not liable for any repercussions arising from the use of its cell biology product(s) in therapeutic/diagnostic or any other non-RUO application(s). 4. abm makes no warranties or representations as to the accuracy of the information on this site. Citations from literature are provided for informational purposes only. abm does not warrant that such information has been shown to be accurate. 5. abm warrants that cell lines shall be viable upon initiation of culture for a period of thirty (30) days after shipment and that they shall meet the specifications on the applicable abm Material Product Information sheet, certificate of analysis, and/or catalog description. Such thirty (30) day period is referred to herein as the "Warranty Period".
Expression Level	CD11c, CD24, MHC-II+, DEC205, Clec9A, B220, CD11b, IRF4, IRF8, CD4
Growth Properties	Adherent, polygonal
Growth Conditions	Use of PriCoat?T25 Flasks (G299) or Applied Cell Extracellular Matrix (G422) is required for cell adhesion to the culture vessels. PriGrow III (TM003) + 10% FBS + 1% Penicillin/Streptomycin Solution (G255), 37.0?, 5% CO?
Quantity	1x106 cells / 1.0 ml
Tissue	Bone
Morphology	Small aggregates
Notes	1. Visually examine the packaging containers for signs of leakage or breakage. 2. Immediately transfer frozen cells from dry ice packaging to a temperature below -130?, preferably in liquid nitrogen vapor phase storage, until ready for use.To ensure the highest level of viability, thaw the vial and initiate culture as soon as possible upon receipt. If continued storage is desired, the vial should only be stored below -130? or in liquid nitrogen vapor phase. Do not store at -70?, as it will result in loss of viability.
Organism	Mouse (M. musculus)
Population Doubling	45 - 55 hours
Species	Mouse (M. musculus)
Propagation	Use of PriCoat T25 Flasks (G299) or Applied Cell Extracellular Matrix (G422) is required for cell adhesion to the culture vessels. Prigrow V (TM015) + 4 mM Glutamax + 10% heat-inactivated FBS + 1% of 7.5% Sodium Bicarbonate Solution + 50 µM beta-mercaptoethanol + 10 mM HEPES (TM058) + 1% Penicillin/Streptomycin Solution (G255), filter the complete media at 0.22 µm before use, 37.0°C, 5% CO?Change the complete media every 2-3 days. Do not let media colour change to yellow.To heat-inactivate FBS:1. Let the FBS bottle completely thaw overnight at 4°C.2. Leave the FBS bottle in water bath for 30 minutes at 56°C.3. Heat-inactivated FBS can be stored at -20°C for long term. Avoid frequent freeze-thaw cycling.To thaw T0528:1. Thaw the vial in 37°C water bath until there is no more than a small cube of ice.2. Transfer the cells to a 15ml tube containing 5ml of pre-warmed complete media.3. Centrifuge the cells at 290xg for 5 minutes.4. Carefully discard supernatant without disturbing the cell pellet and gently resuspend the cells in 1ml of complete media by lightly pipetting up and down.5. Seed the cells at 20,000 - 60,000 cells/cm2.To subculture T0528:1. It is recommended to subculture when cells are at 70-90% confluency.2. Aspirate old media. Some cells in suspension are still viable cells. Alternatively, the supernatant containing the suspended cells can also be collected and centrifuged (Skip to Step 6 in protocol).3. Add 1:1 ratio of 1X sterile PBS and 0.25% Trypsin-EDTA (TM051).4. Incubate cells at room temperature for 3-5 minutes and agitate the culture vessel until 90% of the cells have detached.5. Immediately neutralize the trypsin by adding complete media equal to the volume of trypsin + PBS added.6. Collect the cells and centrifuge at 290xg for 5 minutes.7. Discard the supernatant and gently resuspend the cells in complete media by lightly pipetting up and down.8. Recommended split ratio is no more than 1:3.Stimulation can be performed using PolyIC (5 µg/ml), CpG (2 mM) or LPS (5 µg/ml). These cells are especially sensitive to FBS requirement, thus, it is advised to use the same batch for culturing cells that show best result in supporting their culture. We also recommend the addition of extra 1% HEPES to the complete media to encourage propagation of the cells.To freeze T0528:Recommended freezing medium: Complete growth medium with heat-inactivated FBS to a final concentration of 50% and 5% DMSO.Storage temperature: Liquid nitrogen vapour phase.
Cryopreservation	Cryopreservation Medium (TM024), or complete growth media with 10% DMSO.
Quality Control	1) Direct antigen presentation and cross-antigen presentation were evaluated by the MHC-I (SIINFEKL/OT-I ) and MHC-II (OVA323-339/OT-II) restricted systems; 3) proteome profile and surface markers assessed by RT-PCR; RT-PCR; 3) IL-12 cytokine secretion analyzed using ELISA ; 4) Response to PAMP stimulation evaluated by functional assays

Applications Range	Research Use Only.

Shipping	Ship with dry ice.
Product Format	Frozen
Product Use	This product is intended for laboratory research use only. It is not intended for any animal or human therapeutic use, any human or animal consumption, or any diagnostic use.
Storage	Vapor phase of liquid nitrogen, or below -130?.

Thawing Protocol	1. Thaw cells quickly in a 37? water bath while agitating gently (maximum 2 minutes). The vial cap should be kept above the water level to minimize the risk of contamination. 2. Decontaminate the vial by spraying and wiping the exterior of the vial with 70% ethanol. From this point onwards, all operations should be strictly carried out inside a biological safety cabinet using aseptic conditions. 3. Transfer the cell suspension into a 15ml sterile conical tube containing 5ml of pre-warmed, complete growth media. Centrifuge cells at 125xg for 5-7 minutes. 4. Aspirate the supernatant without disturbing the cell pellet. Re-suspend the cell pellet in the recommended pre-warmed, complete growth media and dispense into a T25 culture flask. 5. Incubate the cells at the recommended conditions.
Subculture Protocol	Volumes given below are for a T75 flask; proportionally increase or decrease the volume as required per culture vessel size. Subculture cells once the culture vessel is 80% confluent. 1. Aspirate the culture media, and add 2-3ml of pre-warmed 0.25% Trypsin-EDTA to the culture vessel. 2. Observe the cells under a microscope to confirm detachment (typically within 2-10 minutes). Cells that are difficult to detach can be put in 37?, for several minutes to facilitate detachment. 3. Neutralize Trypsin-EDTA by adding an equal volume of the complete growth media into the culture vessel. 4. Transfer the culture suspension into a sterile centrifuge tube, and centrifuge at 125xg for 5 minutes. The actual centrifuge duration and speed may vary depending on the cell type. 5. Aspirate the supernatant, and re-suspend the pellet with pre-warmed fresh complete growth media. Add appropriate aliquots of the cell suspension to new culture vessels, as desired. 6. Incubate the cells at the recommended conditions.

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SKU	Supplier No.	Size	Your price	Quantity
BHC10900735	T0227	1x10^6 cells / 1.0 ml	-		Add to Quote

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