BBARL, Inc.

Human Mesenchymal Stem Cells (hMSCs) Serum-Free Medium for cell proliferation suppression

Xyltech™ MSC-01 Xeno-Free
Catalog Number (BBARL/NIPRO): 10401/87-330

100 mL1. Product features
This product is a culture medium suitable for suppressive growth control of human mesenchymal stem
cells (hMSCs). Xyltech™ MSC-01 Xeno-Free can be used to control the growth rate of hMSCs incombination with Xyltech™ Growth MSC, which is a serum-free medium for hMSCs proliferation. Thisproduct is *Xeno-Free culture medium. In addition, there is no need to pre-coat culture vessels with any
type of attachment matrix.
*Xeno-Free: It contains human-derived components. Any other animal-derived component is free.

2. Precautions for use
Xyltech™ MSC-01 Xeno-Free does not contain substances that neutralize trypsin activity. When
subculturing cells with trypsin, it is strongly recommended that the trypsin activity be sufficiently neutralizedwith a trypsin inhibitor. Dilution washing alone does not completely remove trypsin activity and theremaining protease activity will reduce subsequent cell growth.
This product is a research reagent. It cannot be used for human or animal treatment or diagnostic purposes.

3. Storage
Store Xyltech™ MSC-01 Xeno-Free in a freezer (-20°C). After thawing, store in a cool, dark place (2-8°C).
Also, do not refreeze the thawed medium to avoid deterioration of some active ingredients.

4. Thawing and preparation
This product should be thawed at room temperature (20-25°C). Avoid thawing in a water bath. Afterthawing, store the medium in a cool, dark place (2-8°C) and use within one month.
Fine particles may be observed but there is no influence on quality.

5. Example of cell culture protocol for normal human adipose-derived stem cells (ADSCs) cultureusing Xyltech™ MSC-01 Xeno-Free and Xyltech™ Growth MSC (Xeno-Free)
5-1. Cells and reagents・Normal human ADSCs (100 mm-dish)・Xyltech™ MSC-01 Xeno-Free (growth suppression medium)*This product・Xyltech™ Growth MSC Medium(BBARL CAT. #: 10411)
・Xyltech™ Growth MSC Supplement XF(BBARL CAT. #: 10412)
・r-TE (r-Trypsin/EDTA Solution)(NIPRO CAT. #: 87-974)
・s-TI (Synthetic Trypsin Inhibitor Solution)(NIPRO CAT. #: 87-975)
・D-PBS (-)
5-2. Growth control of normal human ADSCs
1. Before using Xyltech™ Growth MSC, add the entire amount of Xyltech™ Growth MSC Supplement
XF (thawed before use) to Xyltech™ Growth MSC Medium and mix well.
2. Warm the Xyltech™ Growth MSC, D-PBS (-), r-TE, and s-TI in a 37°C water bath.
3. Remove the culture supernatant of selected normal human ADSCs that have reached around 80%
confluence (sub confluence).
4. Rinse the cell layer with 5 mL of D-PBS (-).
5. Add 0.5 mL of r-TE and incubate at 37°C for approximately 2 minutes.
6. Add 0.5 mL of s-TI, mix well, gently pipette up and down several times, collect cells from the dish,and centrifuge at 1,000 rpm, for 5 minutes.
7. Aspirate the supernatant and add the appropriate amount of Xyltech™ Growth MSC to resuspend
the cells and seed into a new tissue culture dish.

8. The next day, using a phase-contrast microscope to confirm the cells are engrafted and replace thegrowth medium with the Xyltech™ MSC-01 Xeno-Free culture medium.
9. The cells can be cultured for up to 3 days with Xyltech™ MSC-01 Xeno-Free. The cells start toregrow quickly and become confluent 1-2 days after changing the medium back to the Xyltech™
Growth MSC. Begin subculture and/or experiments with the cells.
5-3. Phase contrast microscope images of normal human ADSCs cultured with Xyltech™ Growth
MSC (Xeno-Fee) or Xyltech™ MSC-01 Xeno-Free

Bars=200 µm

Fig. 1 Normal human ADSCs cultured withXyltech™ Growth MSC (Xeno-Free) for 3days.

Fig. 2 Normal human ADSCs culturedwith Xyltech™ MSC-01 Xeno-Free(growth suppressive medium) for 3 days.

*The protocol is based on experimental results. It may be necessary to adjust seeding density, andpassage timing according to the cells. This protocol is intended for research purposes only.

6. For Inquiries about products
Bourbon Biomedical Advanced Research Laboratories, Inc. (BBARL, Inc.)
1-3-1, Ekimae, Kashiwazaki-city, Niigata,
TEL: +81-257-23-2769 E-mail: support@bourbon-barl.co.jp
URL: https://www.bourbon-barl.co.jp/eg/Copyright©2022 Bourbon Biomedical Advanced Research Laboratories, Inc., All rights reserved.