Immortalized Human Mammary Epithelial Cells (HMEC 2.6) - SV40

Mfr.No.	T0455
Description	Immortalized Human Mammary Epithelial Cells (HMEC 2.6)-SV40 is an immortalized breast epithelial cell line which is non-tumorigenic and do not show anchorage-independent growth. There is a normal distribution of cells in G1, S, and G2/M phase and they express normal breast epithelial markers (E-cadherin, CK7/18, and CK5/14). The cells retain primary characteristics and are valuable for research relating to mammary epithelial biology.
Cell Type	Immortalized Cells
Biosafety Level	2
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".
Donor History	Female
Expression Region	E-cadherin, CK7/18, CK5/14, Neomycin-resistance, G418 and zeocin resistance
Expression Level	K5, K14, vimentin, E-cadherin, p63, K8, K18, K19, CD29, CD49f a-SMA, CD10, Thy-1 when grown in DCFI-1
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 IV (TM004) + 1% FBS (or 35 ?/ml bovine pituitary extract (Hammond)) + 12.5 ng rhEGF (Z100135) + 50 ? fresh ascorbic acid + 2 nM estradiol + 1 ?/ml insulin (TM053) + 2.8 ? hydrocortisone + 0.1 mM ethanolamine + 0.1 mM L-glutamine (G275) + 15 nM sodium selenite + 1% Penicillin/Streptomycin Solution (G255), 37.0?, 5% CO?
Quantity	1x106 cells / 1.0 ml
Tissue	Mammary
Morphology	Cobble-stone\|Spindle shaped
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	Human (H. sapiens)
Population Doubling	24 - 48 hours
Species	Human (H. sapiens)
Propagation	Use of PriCoatTM T25 Flasks (G299) or Applied Cell Extracellular Matrix (G422) is required for cell adhesion to the culture vessels. Grow cells in ECM-coated culture vessels unless otherwise specified in the Propagation Requirements below.Optimized Media:To express basal, luminal, and stem cell markers with some myoepithelial cell markers (a-SMA, CD10, Thy-1), grow cells in DFCI-1 media.To make DFCI-1 the base medium is Prigrow VIII and Prigrow IV (1:1, vol/vol) medium available at abm (TM018 and TM004). To make the complete growth medium, add the following components to the base medium: fetal bovine serum (TM999)* to a final concentration of 1%, Penicillin/Streptomycin Solution (G255) to a final concentration of 1%, HEPES (Gibco) to a final concentration of 10 mM, 12.5 ng/ml Recombinant Human EGF (Z100135) to a final concentration of 12.5 ng/ml, 6.5 ng/ml triiodothyronine (Sigma), 0.545 ng/ml beta-estradiol (Sigma), 1 µg/ml insulin (TM053), 1 µg/ml hydrocortisone (Sigma), 0.006X ethanolamine (Sigma), 14.1 µg/ml phosphoethanolamine (Sigma), 10 µg/ml transferrin (Sigma), 2 mM L-glutamine (G275), 2.6 ng/ml sodium selenite (Sigma), 1 ng/ml cholera toxin (Sigma), 35 µg/ml bovine pituitary extract (Hammond Cell Tech), and 10 µg/ml ascorbic acid (Sigma). Adjust pH to 7.4.Change media every 2-3 days.Carbon dioxide (CO2): 5%, Temperature: 37.0°C.* Do not use heat-inactivated FBS for cell culture unless specified otherwise.For cells to adopt spindle-shape morphology around tight epithelial cell colonies and for the presence of mucin-1 positive cells: grow cells in mammary epithelial growth medium (MEGM; Lonza) supplemented with B27 (10ml/500ml medium), 20 ng/ml Recombinant Human EGF (Z100135), 20 ng/ml Recombinant Human FGF2 (Z101455), 4 µg/ml heparin, and Penicillin/Streptomycin Solution (G255) to a final concentration of 1%.Carbon dioxide (CO2): 6.5%, Temperature: 37.0°C.For luminal differentiation, the absence of myoepithelial differentiation, and the appearance of mucin-1 positive and vimentin-negative cells: grow the cells in DCFI-2 media where the base is Prigrow VIII and Prigrow IV (1:1, vol/vol) medium available at abm (TM018 and TM004). Supplement with 10 mM HEPES (Gibco), 12.5 ng/ml Recombinant Human EGF (Z100135), 6.5 ng/ml triiodothyronine (Sigma), 0.545 ng/ml beta-estradiol (Sigma), 1 µg/ml insulin (TM053), 1 µg/ml hydrocortisone (Sigma), 0.006X ethanolamine (Sigma), 14.1 µg/ml phosphoethanolamine (Sigma), 10 µg/ml transferrin (Sigma), 2 mM L-glutamine (G275), 2.6 ng/ml sodium selenite (Sigma), 1 ng/mg cholera toxin (Sigma), 0.05% bovine serum albumin (BSA), Penicillin/Streptomycin Solution (G255) to a final concentration of 1%, and 10 µg/ml ascorbic acid (Sigma). Adjust pH to 7.4.Carbon dioxide (CO2): 6.5%, Temperature: 37.0°C.
Cryopreservation	Cryopreservation Medium (TM024), or complete growth media with 10% DMSO.
Quality Control	1) Western blot to assess cell lineage-related and stem cell markers; 2) Immunofluorescence staining analysis for lineage markers; 3) Gene expression profiling for gene regulatory pathways

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
BHC10900584	T0455	1x10^6 cells / 1.0 ml	-		Add to Quote

Immortalized Human Mammary Epithelial Cells (HMEC 2.6) - SV40

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