Shreffler:CFSE Labeling

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Line 12: Line 12:
Make '2X' concentration (10 μM) in PBS
Make '2X' concentration (10 μM) in PBS
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*Add 4 mL PBS + 8 μL 5 mM CFSE (for example)
+
*Add 5 mL PBS + 10 μL 5 mM CFSE (for example)
Combine 1:1 PBMC cells + CFSE (e.g. 5 mL PBMCs + 5 mL CFSE) in 15mL tube.
Combine 1:1 PBMC cells + CFSE (e.g. 5 mL PBMCs + 5 mL CFSE) in 15mL tube.

Revision as of 21:45, 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 5 mL PBS + 10 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

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