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General description
Proteolytic enzymes such as trypsin, collagenase and pronase are commonly used for dispersing tissues and cells. These enzymes, however, often injure the cells, are unstable during incubation, can be heterogeneous and also a source of mycoplasma contamination. Dispase-II is used for the preparation of cells from a wide variety of different tissues and organs. Dispase-II has proven to be a rapid and effective, yet gentle, agent for separating many tissues and cells grown in vitro. Dispase is especially suitable for tissue disaggregation and subcultivation procedures since it does not damage cell membranes.
PluriSTEM Dispase-II has been validated to work alongside PluriSTEM Human ES/iPS Medium (SCM130) for the culture and passage of human embryonic and induced pluripotent stem cells.
Dispase-II enzyme is produced in Bacillus polymyxa.
Application
Optimal passaging technique must be determined by end user and varies depending on cell type and culture conditions.
1. Coat 6-well plates with 1:20 Matrigel coating (1.5 ml per well). Incubate at 2-8ºC overnight or coat at RT for at least 30 minutes to 1 hour before use.
2. Remove matrigel coating. Add 2 mL of Complete Media per well.
3. Remove areas of differentiation within culture.
4. Aspirate media.
5. Wash once with 2 mL PBS.
6. Add 1.5 mL Dispase II per well. Incubate for 37ºC for 7 minutes.
7. Wash 2X with 2 mL PBS, w/o Mg and Ca.
8. Add 2 mL Complete Media to each well and use cell scraper to detach colonies.
9. Collect scrapped cells in 15 ml conical tube.
10. Spin 800 rpm for 5 minutes
11. Resuspend in appropriate volume of complete media. Typical splitting ratio is 1:5 – 1:6 depending upon cell density. Cells should be ready for passaging in 5-6 days time using PluriSTEM Human ES/iPS Medium.
Research Category
Stem Cell Research
Research Sub Category
Enzymes & Biochemistry
Physical form
Product is presented in sterile DMEMF12 at 1 mg/ml. Product is filtered through a 0.2 micron filter before freezing.
Dispase-II enzyme is produced in Bacillus polymyxa.
Preparation Note
Store at -20°C for up to 4 months from date of receipt. Once thawed, aliquot in smaller working volumes and store at -20ºC. Avoid multiple freeze thaw cycles to maintain proper enzymatic activity.
Analysis Note
Solution is red with no particulates present. Solution is suitable for cell culture applications.
Legal Information
PLURISTEM is a registered trademark of Merck KGaA, Darmstadt, Germany
Disclaimer
Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
