Immortalized Human Mammary Epithelial Progenitor (K5+/K19+) Cells - hTERT

Mfr.No.	T0452
Description	The immortalized human mammary progenitor cells ?hTERT (K5+/K19+) is a clonal cell population of progenitors coexpressing normal mammary and stem cell markers. It has the ability to self-renew and differentiate into luminal and/or myoepithelial cell lineages. Through propagation in different optimized media, the cells are able to give rise to new population of cells with different morphology and characteristics, such as mucin-1 positive cells, vimentin-negative cells, or expressing basal, luminal, and other stem cell markers. Aldehyde dehydrogenase 1A3 enzyme is noticeably higher in expression, a marker commonly used for isolating normal and tumor mammary stem cells. K5+/K19+ cells have Wnt, Notch, and hedeghog gene regulatory pathways. It is a valuable tool in research in the field of biology, the stem/progenitor origin, and the heterogeneity mechanisms in breast cancer.Western blot was used to assess cell lineage-related and stem cell markers.
Cell Type	Immortalized Cells
Biosafety Level	2
Depositor	University of Nebraska Medical Center
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	a-SMA, CD10, and Thy-1 (with DFCI-1 media conditions). K5, K14, vimentin, E-cadherin, p63, K8, K18, K19, CD29, CD49f
Growth Properties	Adherent, cobblestone \| Spindle-shaped if cultured in mammary epithelial growth media
Growth Conditions	Use of PriCoat?T25 Flasks (G299) or Applied Cell Extracellular Matrix (G422) is required for cell adhesion to the culture vessels. 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 ?/ml insulin (TM053), 1 ?/ml hydrocortisone (Sigma), 0.006X ethanolamine (Sigma), 14.1 ?/ml phosphoethanolamine (Sigma), 10 ?/ml transferrin (Sigma), 2 mM L-glutamine (G275), 2.6 ng/ml sodium selenite (Sigma), 1 ng/ml cholera toxin (Sigma), 35 ?/ml bovine pituitary extract (Hammond Cell Tech), and freshly added 10 ?/ml ascorbic acid (Sigma). Change media every 2-3 days. Carbon dioxide (CO2): 5%, Temperature: 37.0?. * 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 ?/ml heparin, and Penicillin/Streptomycin Solution (G255) to a final concentration of 1%. Carbon dioxide (CO2): 6.5%, Temperature: 37.0?. For information regarding cell differentiation, please refer to the Differentiation Protocol PDF under the Documents Tab.
Quantity	1x106 cells / 1.0 ml
Tissue	Mammary
Morphology	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	57 - 67 hours
Species	Human (H. sapiens)
Propagation	Use of PriCoatTMT25 Flasks (G299) or Applied Cell Extracellular Matrix (G422) is required for cell adhesion to the culture vessels. Grow cells inECM-coated culture vessels unless otherwise specified in the Propagation Requirements below.The base medium for this cell line is Prigrow III medium available at abm, Cat. No. TM003. To make the complete growth medium, add the following components to the base medium: fetal bovine serum (TM999)* to a final concentration of 10%, L-glutamine (G275) to a final concentration of 1%, and Penicillin/Streptomycin Solution (G255) to a final concentration of 1%.Change media every 2-3 days.Carbon dioxide (CO2): 5%, Temperature: 33.5°C.* Do not use heat-inactivated FBS for cell culture unless specified otherwise.
Cryopreservation	Cryopreservation Medium (TM024), or complete growth media with 10% DMSO.
Quality Control	1.) Telomerase-detection assay was used to detect telomerase activity. 2.) Flow cytometry confirmed expression of T antigen.

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
BHC10900583	T0452	1x10^6 cells / 1.0 ml	-		Add to Quote

Immortalized Human Mammary Epithelial Progenitor (K5+/K19+) Cells - hTERT

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