Poly-L-ornithine-coated plates

MATERIALS

Ethanol (70%) (if coating 24-well plates)

Phosphate-buffered saline (PBS; 1X, sterile)

Poly-L-ornithine hydrochloride (Sigma P2533)

Coverslips (12-mm; Bellco Glass 1943-10012) (if coating 24-well plates)

Foil (optional; see Step 2.iii)

Hood, tissue culture (with UV light)

Plates (six- or 24-well)

1. Prepare 1.5-mg/mL (100X) aliquots of poly-L-ornithine hydrochloride by diluting in H₂O. Store at −20°C.
2. Prepare coated plates:
- To coat 6-well plates for biochemistry:
- i. Dilute the 100X poly-L-ornithine to 1X in sterile 1X PBS, and pipette enough of the solution to cover the surface of each well. Incubate plates overnight at 37°C.
- ii. The next day, aspirate the coating solution and wash three times with H₂O, making sure the H₂O thoroughly fills each well. Air dry in a tissue culture hood until no H₂O remains.
- iii. Store covered in foil at 4°C if the plate is not to be used immediately.
- To coat 24-well plates for morphology assays:
- iv. Sterilize a packet of 12-mm coverslips by rinsing for several hours in 70% ethanol and drying overnight under the UV light of the tissue culture hood.
- v. Place one coverslip in each well of the 24-well plate, add 0.5 mL of 1X poly-L-ornithine coating solution (diluted as in Step 2.i) per well, and follow the incubation/rinsing procedure described in Step 2.ii, using at least 1 mL of H₂O per well.