I cut fresh adipose tissue into pieces using a scalpel. After that I incubated tissue with 1.5mg/ml collagenase at 37 degrees for 12 hours on a rotor. I then passed tissue/collagenase solution through a 70um cell strainer and centrifugated the flow-through at 500g for 10 minutes. I didn't see the adipose ring at the top and the SVF pellets at the bottom after centrifuge. Instead, a thick lipid layer was left on the strainer.
I was wondering if the cells are killed due to the long digestion time or the small strainer size blocked the flow through of lipid and cells (Some researchers used 200um strainer).
You can benefit from the following links.
1. Orr, J.S., Kennedy, A.J., Hasty, A.H. Isolation of Adipose Tissue Immune Cells. J. Vis. Exp. (75), e50707, doi:10.3791/50707 (2013).
2. Swathi SundarRaj, Abhijeet Deshmukh, Nancy Priya, Vidya S. Krishnan, Murali Cherat, and Anish Sen Majumdar, “Development of a System and Method for Automated Isolation of Stromal Vascular Fraction from Adipose Tissue Lipoaspirate,” Stem Cells International, vol. 2015, Article ID 109353, 11 pages, 2015. https://doi.org/10.1155/2015/109353.
3. Li-TingLiaKuang-TaYao,Shou-ChengTeng, Tiffany P.Sun Ching-KuoChenaChia-ChunChenaMing-LunHsu.Injectable adipose tissue combined with stem cells for soft-tissue augmentation: A pilot study for dental applications. 11, (4), December 2016, Pages 377-386
4. Isolation of Stromal Stem Cells from Human Adipose Tissue. www.collaslab.com
Thanks Entedhar for sharing some relevant articles, especially for the 3rd one which optimized the digestion time, centrifuge force and time. I will try 30 to 90 min for digestion and 1200g for 3mins for centrifuge.
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