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Mouse Embryonic Blasticidin and Zeocin-resistant (MMMbz) Immortalized Feeder Cells

SKU : BHC10901970

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Applied Biological Materials (abm) Inc.

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Mfr.No.	T2028
Description	Supporting cells, such as short-term cultures of fibroblasts, are used to cultivate human embryonic stem (ES) cells, however they are susceptible to senescence after a number of passages. The Mouse Embryonic Blasticidin and Zeocin-resistant (MMMbz) Immortalized Feeder Cells eliminates the problems associated with the use of primary cell lines. This cell line has a doubling time of 16 hours and may be used indefinitely to propagate human embryonic stem cells. It has the ability to facilitate the growth of hES H9 and hES H1. MMMbz cells are resistant to blasticidin and zeocin due to transgene expression allowing for selection of hES cells transfected with genes conferring resistance.
Cell Type	Stable Cell Lines
Biosafety Level	2
Depositor	Harvard University
Disclaimer	1. Sale of this item is subjected to the completion of a Material Transfer Agreement (MTA) by the purchasing individual/institution for each cell line. If you have any questions regarding this, please contact us at licensing@abmgood.com. 2. 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. 3. 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. 4. 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). 5. 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. 6. 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".
Donor History	Fetal
Growth Properties	Adherent, epithelial-like
Growth Conditions	Use of PriCoat?T25 ECM-coated flasks (G299) or Applied Cell Extracellular Matrix (G422) is recommended for cell adhesion to the culture vessels. PriGrow III (TM003) + 10% FBS + 1% Penicillin/Streptomycin Solution (G255), 37.0?, 5% CO?. Do not use heat-inactivated serum for cell culture unless specified otherwise. To support the growth of hES cells:?rradiate MMMbz cells with 6000 rads and seed at a density of 1.3x105/cm2?n propagation medium. The next day add hES in Knockout DMEM (Invitrogen), supplemented with 20% Knockout Serum Replacement (Invitrogen), 1mM L-glutamine (G275), 1% MEM non-essential amino acids (Invitrogen), 0.1mM 2-mercaptoethanol (CH045) and bFGF at 4 ng/ml (Z200015).
Quantity	10⁶ cells/1.0 ml
Tissue	Embryo
Morphology	Fibroblast-like
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)
Species	Mouse (M. musculus)
Propagation	Grow cells in T25 ECM-coated flasks (G299) with the following conditions. The base medium for this cell line is Prigrow III medium available at abm, Cat. No. TM003. To make the completed growth medium, add the following components to the base medium: Bovine Calf Serum (available from Hyclone, SH30072) to final concentration of 10% and Penicillin/Streptomycin Solution (G255) to a final concentration of 1%. Carbon dioxide (CO2): 5%, Temperature: 37.0°C. To freeze: suspend cells in PriGrow III (TM003) with 10% bovine calf serum and 10% sterile DMSO. To support the growth of hES cells: irradiate MMMbz cells with 6000 rads and seed at a density of 1.3x105/cm2 in propagation medium. The next day add hES in Knockout DMEM (Invitrogen), supplemented with 20% Knockout Serum Replacement (Invitrogen), 1mM L-glutamine (G275), 1% MEM non-essential amino acids (Invitrogen), 0.1mM 2-mercaptoethanol (CH045) and bFGF at 4 ng/ml (Z200015).
Cryopreservation	Cryopreservation Medium (TM024), or complete growth media with 10% DMSO.
Source Organ	Reproductive
Quality Control	Selective growth with media containing blasticidin and zeocin to select for cells with transgene expression.

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
BHC10901970	T2028	1x10^6 cells / 1.0 ml	-		Add to Quote

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