ATCC ATCC Logo 0
  • Quick Order
  • Careers
  • Support

Adipocyte Differentiation Toolkit for Adipose-Derived MSCs and Preadipocytes

PCS-500-050

The Adipocyte Differentiation Toolkit for Adipose-derived MSCs and Preadipocytes contains medium and reagents designed to induce adipogenesis in actively proliferating Adipose-Derived Mesenchymal Stem Cells and Preadipocytes with high efficiency, and to support the maturation of derived adipocytes during lipid accumulation.
Product category
Reagents
Applications
Stem cell research
Cell growth and viability
Cell and gene therapy (CGT) development
Product format
Frozen
Shipping information
1 kit
Storage conditions
-20°C or colder, -70°C for long-term storage

Documentation

ATCC determines the biosafety level of a material based on our risk assessment as guided by the current edition of Biosafety in Microbiological and Biomedical Laboratories (BMBL), U.S. Department of Health and Human Services. It is your responsibility to understand the hazards associated with the material per your organization’s policies and procedures as well as any other applicable regulations as enforced by your local or national agencies.

Detailed product information

General

Specific applications
The Adipocyte Differentiation Toolkit for Adipose-derived MSCs and Preadipocytes contains medium and reagents designed to induce adipogenesis in actively proliferating Adipose-Derived Mesenchymal Stem Cells and Preadipocytes with high efficiency, and to support maturation of derived adipocytes during lipid accumulation. The Toolkit is composed of the following components:
  1. One vial containing 1 mL of AD Supplement for use in initiation of adipocyte differentiation
  2. One bottle containing 5 mL of ADM Supplement for use in maintaining adipocyte differentiation
  3. One bottle containing 100 mL of Adipocyte Basal Medium for use in both initiation and maintenance of adipocyte differentiation

*The Adipocyte Differentiation Toolkit for Adipose-derived MSCs and Preadipocytes provides enough medium and reagents for differentiation of ~6.8 x 105 cells when plated at a recommended density of 18,000 viable cells/cm2 in 4 wells of a 6 well tissue culture format. 



Handling information

Handling procedure
Unpacking and Storage Instructions

  1. Check all containers for leakage or breakage.
  2. Store the differentiation toolkit at -20°C in a freezer that is not self-defrosting. Do not refreeze the supplements once thawed.
  3. If thawed upon arrival, the supplements can be stored at 2 to 8°C as long as they are added to the Adipocyte Basal Medium within 72 hours.
  4. Once prepared, both supplemented media are stable for up to three weeks when stored in the dark at 2 to 8°C.
Note: Instructions for preparing (1) the Initiation Medium with AD Supplement, and (2) the Maintenance Medium with the ADM Supplement are provided below. Please see the “Adipocyte Differentiation Media Preparation” section before proceeding.

Antimicrobials and phenol red are not required but may be added to the 100 mL bottle of Adipocyte Basal Medium if desired prior to supplementation. The recommended volume of each optional component to be added to the Adipocyte Basal Medium is summarized in Table 1.

Component

Volume

Final Concentration

Gentamicin-Amphotericin B Solution

0.1 mL

Gentamicin: 10 µg/mL

Amphotericin B: 0.25 µg/mL

Penicillin-Streptomycin-Amphotericin B Solution

0.1 mL

Penicillin: 10 Units/mL

Streptomycin: 10 µg/mL

Amphotericin B: 25 ng/mL

Phenol Red

0.1 mL

33 µM


Preparing Cells for Adipocyte Differentiation

  1. Follow the instructions for the growth of Adipose-Derived Mesenchymal Stem Cells (ATCC PCS-500-011). It is recommended that the cells not be passaged more than four (4) times before initiating adipocyte differentiation.
  2. When cells are 70-80% confluent, passage them into a tissue culture plate at a density of 18,000 cells/cm2. Adjust the number of cells and volume of media according to the tissue culture plate used.
  3. Example: For a 6 well tissue culture plate with a surface area of 9.5 cm2/well, add a total of 171,000 viable cells to each well containing 2 mL of Mesenchymal Stem Cell Basal Medium (ATCC PCS-500-030) supplemented with Mesenchymal Stem Cell Growth Kit–Low Serum (ATCC PCS-500-040) components.
  4. Gently rock the plate back and forth and side to side to evenly distribute cells before incubation. Do not swirl.
  5. Incubate the cells at 37°C with 5% CO2 for 48 hours before initiating adipocyte differentiation.

Adipocyte Differentiation Media Preparation

The adipocyte differentiation process requires two separate media preparations: one for initiation and one for maintenance. Stock solutions of these media can be prepared in tandem in advance as follows: 

  1. Thaw all three  components of the differentiation kit and warm to 37°C in a water bath. Note: It may be necessary to shake the AD Supplement and the ADM Supplement upon warming to help re-dissolve any components that may have precipitated out of solution upon freezing.
  2. Decontaminate the external surfaces of all three kit components by spraying them with 70% ethanol.
  3. Using aseptic technique and working in a laminar flow hood or biosafety cabinet:
    1. Transfer 15 mL of Adipocyte Basal Medium and 1 mL of AD Supplement to a sterile 50 mL conical tube, using a separate sterile pipette for each transfer. This is your working stock of Adipocyte Differentiation Initiation Medium used during the first 96 hours of differentiation.
    2. Add 5 mL of ADM Supplement to the remaining 85 mL of Adipocyte Basal Medium. This is your working stock of Adipocyte Differentiation Maintenance Medium.
  4. Tightly cap the each container of media and swirl the contents gently to assure a homogeneous solution. Do not shake forcefully to avoid foaming. Label and date the bottle.
  5. Each container of differentiation medium should be stored in the dark at 2°C to 8°C (do not freeze). When stored under these conditions, the differentiation media is stable for up to three weeks.

Adipocyte Differentiation Procedure

Initiation Phase

  1. After incubating the prepared Adipose-Derived Mesenchymal Stem Cells for (as described above), carefully aspirate the media from the wells.
  2. Immediately rinse the cells once by adding 2 mL of room-temperature D-PBS (ATCC® 30-2200) to each well, then carefully aspirate the PBS from the wells.
  3. Add 2 mL of pre-warmed (37°C) Adipocyte Differentiation Initiation Medium to each well to begin the adipocyte differentiation process. Note: It is recommended that you transfer the required volume of media to a sterile tube for pre-warming prior to each feeding rather than repeatedly re-warming the entire working stock.
  4. Incubate the cells at 37°C with 5% CO2 for 48 hours.
  5. Feed the cells by carefully removing half the volume of media (1 mL) from each well and adding another 2 mL of pre-warmed (37°C) Adipocyte Differentiation Initiation Medium to each well. Important: DO NOT TILT plate during aspiration. It is important that the cell monolayer is not exposed to air during this and subsequent steps to ensure that developing lipid vesicles do not burst.

Maintenance Phase

  1. Incubate the cells at 37°C with 5% CO2 for 48 hours.
  2. Carefully remove 2 mL of media from each well (leaving 1 mL) and replace with 2 mL of pre-warmed (37°C) Adipocyte Differentiation Maintenance Medium in each well. Important: DO NOT TILT plate during aspiration. It is important that the cell monolayer is not exposed to air during this and subsequent steps to ensure that developing lipid vesicles do not burst.
  3. Repeat Steps 6 and 7 every 3-4 days for another 11 days until adipocytes reach full maturity.  (Full maturity will be reached 15 days after the beginning of initiation phase, or 17 days from initial plating of cells.)
  4. Cells can be used at any phase of adipocyte differentiation as predicated upon experimental design.  To confirm lipid accumulation, cells can be fixed and stained with Oil Red O.
Cryopreservation
1. Check all containers for leakage or breakage.

2. Store the differentiation toolkit at -20°C in a freezer that is not self-defrosting. Do not refreeze the supplements once thawed.

3. If thawed upon arrival, the supplements can be stored at 2°C-8°C as long as they are added to the Adipocyte Basal Medium within 72 hours.

4. Once prepared, both supplemented media are stable for up to three weeks when stored in the dark at 2°C-8°C.

Note: Instructions for preparing (1) the Initiation Medium with AD Supplement, and (2) the Maintenance Medium with the ADM Supplement are provided below. Please see the “Adipocyte Differentiation Media Preparation” section before proceeding.

Quality control specifications

Mycoplasma contamination
Not detected
Functional tests
Differentiation of cells into adipocytes as demonstrated by Oil Red O staining.
A Certificate of Analysis (COA) is available upon request for each lot

Legal disclaimers

Intended 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.
Warranty

The product is provided 'AS IS' and the viability of ATCC® products is warranted for 30 days from the date of shipment, provided that the customer has stored and handled the product according to the information included on the product information sheet, website, and Certificate of Analysis. For living cultures, ATCC lists the media formulation and reagents that have been found to be effective for the product. While other unspecified media and reagents may also produce satisfactory results, a change in the ATCC and/or depositor-recommended protocols may affect the recovery, growth, and/or function of the product. If an alternative medium formulation or reagent is used, the ATCC warranty for viability is no longer valid.  Except as expressly set forth herein, no other warranties of any kind are provided, express or implied, including, but not limited to, any implied warranties of merchantability, fitness for a particular purpose, manufacture according to cGMP standards, typicality, safety, accuracy, and/or noninfringement.

Disclaimers

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. Any proposed commercial use is prohibited without a license from ATCC.

While ATCC uses reasonable efforts to include accurate and up-to-date information on this product sheet, ATCC makes no warranties or representations as to its accuracy. Citations from scientific literature and patents are provided for informational purposes only. ATCC does not warrant that such information has been confirmed to be accurate or complete and the customer bears the sole responsibility of confirming the accuracy and completeness of any such information.

This product is sent on the condition that the customer is responsible for and assumes all risk and responsibility in connection with the receipt, handling, storage, disposal, and use of the ATCC product including without limitation taking all appropriate safety and handling precautions to minimize health or environmental risk. As a condition of receiving the material, the customer agrees that any activity undertaken with the ATCC product and any progeny or modifications will be conducted in compliance with all applicable laws, regulations, and guidelines. This product is provided 'AS IS' with no representations or warranties whatsoever except as expressly set forth herein and in no event shall ATCC, its parents, subsidiaries, directors, officers, agents, employees, assigns, successors, and affiliates be liable for indirect, special, incidental, or consequential damages of any kind in connection with or arising out of the customer's use of the product. While reasonable effort is made to ensure authenticity and reliability of materials on deposit, ATCC is not liable for damages arising from the misidentification or misrepresentation of such materials.

Please see the material transfer agreement (MTA) for further details regarding the use of this product. The MTA is available at www.atcc.org.

Frequently Asked Questions

References

Need assistance with this product? Contact our Technical Support team.

Telephone

US and Puerto Rico
800-638-6597

Outside the US
+1-703-365-2700

Hours of Operation

Monday-Friday
9:00am - 5:00pm
US Eastern Time