ASC/TERT3002021-10-13T17:50:28+00:00

Adipose tissue

ASC/TERT300

Evercyte ́s ASC/TERT300 cell line has been developed from adipose-derived mesenchymal stem cells, can be grown without limitations, shows typical markers and functions of MSCs and therefore represents a valuable model for studying processes such as differentiation, inflammation, tissue homeostasis and repair. In recent years evidence has accumulated that the potency of mesenchymal stem cells at least in part is mediated by secreted extracellular vesicles (EVs). ASC/TERT300 cell line has been established under conditions that will allow production of EVs for clinical applications.

General information

Cat#: CHT-001-0300

Organism: homo sapiens
Tissue, cell type: adipose tissue, mesenchymal stem cells
Morphology:
mesenchymal, spindle-shaped morphology
Life span extension:
ectopic expression of hTERT
Quality:
free from contaminations (bacteria incl. mycoplasma, fungi, HIV, HAV, HBV, HCV, Parvo-B19) and cross-contaminations

Morphology and growth characteristics

ASC/TERT300 cells can be grown for a minimum of 60 population doublings with a stable growth rate and without showing signs of growth retardation. The cells are characterized by the typical spindle-shaped mesenchymal morphology.

Expression of MSC marker proteins CD73, CD90, CD105

ASC/TERT300 cells homogenously express typical marker proteins of mesenchymal stem cells such as CD73, CD90 and CD105, whereas the hematopoietic stem cell marker CD34 is not expressed (green peaks). Cells stained with isotype control antibodies are used as negative control (red peaks).

Differentiation potential towards adipocytes, osteoblasts and chondrocytes

ASC/TERT300 cells can be induced to differentiate towards adipocytes (oil red O staining), osteoblasts (Alizarin red staining) and chondrocytes ( alcian blue staining).

FAQs

What is the difference between ASC/TERT1 and ASC/TERT300 cells?2021-10-13T17:46:35+00:00

Both cell lines originate from adipose tissue. While ASC/TERT300 was manufactured under conditions that will allow later use for production of GMP grade extracellular vesicles, ASC/TERT1 is used exclusively for scientific purpose.

Can the cells be used for production of clinical grade extracellular vesicles?2021-10-11T14:56:57+00:00

Yes, a cell bank was established that can be transferred to a GMP facility for establishment of a production master cell bank. The cell line was established under xeno-free conditions with full documentation of any manipulation step.

In vitro propagation

Mesencult – ACF Plus Culture Kit

Mesencult – ACF Plus Culture Kit including Animal Component-Free Cell Attachment (STEMCELL TECHNOLOGIES, Cat# 05448)

2 mM GlutaMAX-I (Gibco, Cat# 35050-038)

200 µg/ml G418 (InvivoGen, Cat# ant-gn5)

Additional material & reagents

Phosphate buffered saline (PBS) (Sigma, Cat# D8537)

CTS TrypLE Select Enzyme (Gibco, Cat# A1285901)

Protocol passaging of ASC/TERT300
Passaging of cells

The new culture flasks have to be pre-coated with ACF Cell Attachment Substrate following the instructions of the manufacturer. Briefly, dilute the substrate 1:300 in PBS and transfer the diluted substrate to the culture flasks (72 µl/cm²), incubate at least 2 hours at room temperature.

Before introducing cells, remove excess of ACF substrate and rinse flask once with PBS (160 µl/cm²).
Then, add CTS TrypLE Select Enzyme solution (20 µl/cm²), make sure that all cells have been in contact with this solution and incubate the culture flask at 37°C for approximately 2-3 min.
Observe cell detachment under an inverted microscope. As soon as all cells are detached (if necessary agitate the cells by gently hitting the flask), add growth medium (about 160 µl/cm²) and centrifuge at 300 g for 5 min
Discard the supernatant, resuspend the cell pellet in the remaining droplet and add growth medium (about 160 µl/cm²).

Then, transfer appropriate aliquots of the cell suspension to new culture vessels supplemented with growth medium (final volume of 240 µl/cm²).
A split ratio of 1:6 – 1:8 twice a week is recommended (after cells have reached about 70-80 % confluence).

Cultivate cells at 37°C in a humidified atmosphere with 5% CO2.

Cryopreservation

Freezing medium

CryoStor(R) cell cryopreservation medium CS10 (Sigma Aldrich, Cat# C2874

Additional material & reagents

Phosphate buffered saline (PBS) (Sigma, Cat# D8537)
CTS TrypLE Select Enzym (Gibco, Cat# A1285901)

Protocol cryopreservation of ASC/TERT300
Freezing of cells

Detach the cells after having reached about 70-80 % confluence from the culture vessel by using CTS TrypLE Select Enzym solution (Protocol passaging of ASC/TERT300).

Resuspend the detached cells in growth medium and centrifuge at 300 g for 5 min.

Discard the supernatant, resuspend the resulting cell pellet in the remaining droplet and add freezing medium (tempered to 4°C) to reach a cell density of about 3-5 x 105 cells/ml (for thawing in a 25 cm² culture flask).
Add 1 ml of this cell suspension to each pre-cooled cryovial and immediately transfer the cells to -80°C.

After 24 hours transfer the vials to the liquid nitrogen tank.
Thawing of cells

Original Evercyte cells are to be thawed in a T25 roux flask

Pre-coat a cm² roux flask with ACF Cell Attachment substrate (Protocol passaging of ASC/TERT300).

Add 6 ml of complete growth medium to a pre-coated 25 cm² culture flask and place the culture flask in the incubator for at least 30 min to allow the medium to reach its normal pH.
Take a vial of frozen cells, rinse it outside with Ethanol and pre-warm in the hand until one last piece of frozen cells is seen.
Then, immediately transfer the content of the vial to a 15 ml centrifugation tube pre-filled with 9 ml of medium pre-cooled to 4°C and centrifuge for 5 min at 170 g.

Discard the supernatant and resuspend the cell pellet in the remaining droplet.
Add 1 ml of the pre-warmed medium to the cells, transfer them to the prepared culture flask and incubate at 37°C in a suitable incubator.
Passage the cells the day after thawing.

Protocol passaging of ASC/TERT300

Product data sheet – certificate of analysis

Product data sheet (PDS) download
Certificate of analysis (CoA) is available upon request | please contact us indicating the respective LOT number

Protocols

Preparation of ASC/TERT300 cell growth medium
Protocol passaging of ASC/TERT300
Protocol cryopreservation of ASC/TERT300

Data on Markers and Functions

Morphology and growth characteristics
Expression of MSC marker proteins CD73, CD90, CD105
Differentiation potential towards mesenchymal lineages

Licence Conditions

The business concept of Evercyte is to out-license telomerized cells to our customers. The license conditions depend on whether the contract partner is a for profit or a nonprofit organization and the intended use of the cells.

Nonprofit organizations

Evercyte grants licenses for an unlimited period to academic or nonprofit-organizations, whereby the use of Evercyte cell lines is restricted to research & development purposes and non-commercial use. The cells are not intended for human use.
The customers have to agree to the conditions described in our material transfer agreement as well as accept our general terms and conditions.
On time payment for unlimited use: EUR 1300

Profit organizations

Pharmaceutical – chemical- cosmetic industries
Evercyte grants licenses for commercial organizations, whereby we offer an initial testing phase for a flat fee that allows our customers to test our cells in their laboratories for a period of 6 months.
Thereafter, annual license fees fall due, depending on the cell line of interest. Besides offering cell lines for research & development purposes, we also have established cell factories that qualify for production of clinical grade extracellular vesicles for human application.
The customer has to agree to the conditions described in our license agreements.
Contract research organizations (CRO)
Evercyte grants licenses for contract research organizations, whereby we offer an initial testing phase for a flat fee that allows our customers to test our cells in their laboratories. Thereafter, we would negotiate a royalty based long-term license agreement individually.
The use of the cells during these phases is restricted to research & development purposes. The cells are not intended for human use. The customers have to agree to the conditions described in our material transfer agreement and accept our general terms and conditions.
Initial license fee for 6 months: EUR 2000
Annual license fee R&D: royalty based
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Customer Reviews

“I have had the pleasure of working with Evercyte for the last few years. We continually rely on Evercyte because of the high-quality data that they produce, their diligent responsiveness, and their excellent customer service.”

 

Josh Garlich, Senior Research Scientist, Apellis Pharmaceuticals, Inc.

“Cytonus has been working with Evercyte from many years as they are a trusted partner and have always delivered the highest quality cell lines to advance our platform.  We routinely draw on their expertise to meet cellular engineering challenges and they have not disappointed.”

Remo Moomiaie-Qajar, Cytonus Therapeutics, Inc.

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Customer Reviews

“I have had the pleasure of working with Evercyte for the last few years. We continually rely on Evercyte because of the high-quality data that they produce, their diligent responsiveness, and their excellent customer service.”

 

Josh Garlich, Senior Research Scientist, Apellis Pharmaceuticals, Inc.

“Cytonus has been working with Evercyte from many years as they are a trusted partner and have always delivered the highest quality cell lines to advance our platform.  We routinely draw on their expertise to meet cellular engineering challenges and they have not disappointed.”

Remo Moomiaie-Qajar, Cytonus Therapeutics, Inc.

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WJ-MSC/TERT273
Cat#: CHT-021-0273

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