Shreffler:CFSE Labeling: Difference between revisions
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===Materials=== | ===Materials=== | ||
5,6-CFDA/SE [http://products.invitrogen.com:80/ivgn/en/US/adirect/invitrogen?cmd=catProductDetail&entryPoint=adirect&productID=C1157&messageType=catProductDetail Invitrogen C1157] dissolved in DMSO at 5 mM and stored at - | 5,6-CFDA/SE [http://products.invitrogen.com:80/ivgn/en/US/adirect/invitrogen?cmd=catProductDetail&entryPoint=adirect&productID=C1157&messageType=catProductDetail Invitrogen C1157] dissolved in DMSO at 5 mM and stored at -20°C. | ||
===Procedure=== | ===Procedure=== | ||
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Make '2X' concentration (10 μM) in PBS | Make '2X' concentration (10 μM) in PBS | ||
*Add | *Add 4 mL PBS + 8 μL 5 mM CFSE (for example) | ||
Combine 1:1 PBMC cells + CFSE ( | Combine 1:1 PBMC cells + CFSE (e.g. 5 mL PBMCs + 5 mL CFSE) in 15mL tube. | ||
Place in H<sub>2</sub>O bath x 10 min. | Place in 37°C H<sub>2</sub>O bath x 10 min. | ||
Wash in 10 mL complete medium | Wash in 10 mL complete medium. | ||
Resuspend in medium at desired density | Resuspend in medium at desired density | ||
[[Category:Protocol]] | [[Category:Protocol]] | ||
Revision as of 18:44, 19 May 2009
Materials
5,6-CFDA/SE Invitrogen C1157 dissolved in DMSO at 5 mM and stored at -20°C.
Procedure
Suspend PBMCs at 10x106 cells/mL in PBS alone.
Cells should be in PBS for CFSE labeling.
If CFSE negative control is needed, remove cells now.
Make '2X' concentration (10 μM) in PBS
- Add 4 mL PBS + 8 μL 5 mM CFSE (for example)
Combine 1:1 PBMC cells + CFSE (e.g. 5 mL PBMCs + 5 mL CFSE) in 15mL tube.
Place in 37°C H2O bath x 10 min.
Wash in 10 mL complete medium.
Resuspend in medium at desired density
