Cytospinning Cells on Slides
Poly-L (or Poly-D) Lysine Coating Slides
- Prepare 1 mg/mL stock of Poly-L lysine in ddH2O
- Dilute 1:100 (CF 10 mg/mL) in ddH2O
- Coat standard glass slides by soaking in solution for 1 hour
- Air dry for 1 hour
- Slides are now ready for use, or store dry at 4oC
Cytospining:
- Prepare cells in media or PBS, cell number range 5K-50K in 400-800 ml (suggest 25 K in 500 ml).
- Prepare chamber: insert slide into frame with Poly-L lysine coating up, insert gasket, and clamp with clips on either side.
- Insert slide chamber into cytospin with the slide facing outward.
- Load cells into chamber.
- Spin, standard cells spin speed 10, time 4 minutes. Use higher spin speed and longer time for small cells (e.g. spin speed 13, time 6 minutes).
- Aspirate media. Fix cells with 4% PFA by adding equal volume used for loading cells.
- Spin again.
- Wash 2X with PBS (no spinning required).
- Take apart chamber. Use pap pen to draw square around the cells.
- Add additional PBS to cover cells. (Never let them dry!) Store in a humidity chamber at 4oC until use.
