Shreffler:CFSE Labeling: Difference between revisions
From OpenWetWare
Jump to navigationJump to search
(Undo revision 313541 by Wayne G. Shreffler (Talk)) |
|||
| (7 intermediate revisions by 2 users not shown) | |||
| Line 28: | Line 28: | ||
#Lyons pmid=8176234 | #Lyons pmid=8176234 | ||
</biblio> | </biblio> | ||
==Contact== | ==Contact== | ||
*Who has experience with this protocol? | *Who has experience with this protocol? | ||
*[[Special:Emailuser/Wayne G. Shreffler|Email Wayne Shreffler through OpenWetWare]] | *[[Special:Emailuser/Wayne G. Shreffler|Email Wayne Shreffler through OpenWetWare]] | ||
[[Category:ShreffLab]] | |||
[[Category:Protocol]] | |||
[[Category:Flow cytometry]] | |||
[[Category:Immunology]] | |||
Latest revision as of 11:23, 10 June 2009
Overview
CFSE is used to fluorescently label live cells in order to track them. 5,6-CFDA/SE is membrane permeable, but trapped in the cell when converted by intracellular esterases. CFSE is equally partitioned to daughter cells during division and can be used to measure cell proliferation.
Materials
- 15 mL polypropylene tubes
- PBS
- 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.
- Ensure that cells are uniformly suspended when CFSE is added.
- If CFSE negative control is needed, remove cells now.
- Make '2X' concentration (10 μM) in PBS
- For example: add 5 mL PBS + 10 μL 5 mM CFSE
- Combine 1:1 PBMC cells + CFSE (e.g. 5 mL PBMCs + 5 mL CFSE) in 15mL tube and mix completely.
- Place in 37°C H2O bath x 10 min.
- Wash in 10 mL complete medium.
- Resuspend in medium at desired density
Discussion
References
- Lyons AB and Parish CR. Determination of lymphocyte division by flow cytometry. J Immunol Methods. 1994 May 2;171(1):131-7. DOI:10.1016/0022-1759(94)90236-4 |
Contact
- Who has experience with this protocol?
- Email Wayne Shreffler through OpenWetWare
