Difference between revisions of "FM4-64 Labeling of Yeast Cells"
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==Materials== | ==Materials== | ||
| − | *Instead of FM4-64 we use SynaptoRed (Sigma #S6689) | + | *Instead of FM4-64 we use SynaptoRed (Sigma #S6689). This is resuspended 1mg to 500 uL (2ug/mL) with DMSO. |
*Media (see [[YPD Media and Agar]]) | *Media (see [[YPD Media and Agar]]) | ||
*Dye solution. For each cell line need 250 uL media + 3 uL dye. | *Dye solution. For each cell line need 250 uL media + 3 uL dye. | ||
Revision as of 19:40, 7 December 2009
Materials
- Instead of FM4-64 we use SynaptoRed (Sigma #S6689). This is resuspended 1mg to 500 uL (2ug/mL) with DMSO.
- Media (see YPD Media and Agar)
- Dye solution. For each cell line need 250 uL media + 3 uL dye.
Protocol
- Grow and overnight culture of yeast in 4 mL of the appropriate media.
- Spin cells down and resuspend in dye solution FM4-64. Wrap in alluminum foil and incubate with shaking at 24C for 1h.
- Spin down cells and wash 2x with media.
- Add cells to 5 mL media and mix. Take a 1 mL sample and measure A600
- Incubate 3h with shaking at 24C. Take a 1 mL sample and measure A600. This should be one full doubling from the previous reading.
- Mount cells on coverslip and analyze by fluorescent microscopy
