Adipocyte Differentiation Stain (Oil Red O)

Oil Red O staining

  1. Rinse cells two times with PBS
  2. Fix cells in 1mL/6-well 10% formalin/PBS for one hour (no more than 2 hours) at RT.
  3. Rinse plates two times with ddH2O
  4. Stain with 1mL/6-well Oil Red O (stain prep below) for 2 hours at RT.
  5. Aspirate stain and wash cells gently with water a many times (around 15-20, until background stain comes off).
  6. Stained fat droplets will be dark brown/red in color.
  7. Leave some ddH2O on the plates and image.
  8. Either keep moist with PBS or air dry.

Preparation of Oil Red O:

  1. 1.05g Oil Red O dissolved in 300mL isopropanol and leave overnight at RT w/o stirring
  2. Filter with 0.45um membrane, add 250mL ddH2O and leave solution overnight at 4°C without stirring
  3. Filter 1x with 0.45um filter
  4. Filter 1x with 0.22um filter
  5. store at RT