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crystal violet assay

Crystal Violet Assay

This is a simple assay useful for obtaining quantitative information about the relative density of cells adhering to multi-well cluster dishes. The dye in this assay, crystal violet, stains DNA. Upon solubilization, the amount of dye taken up by the monolayer can be quantitated in a spectrophotometer or plate reader.

  1. Carefully remove culture medium from wells.
  2. Wash plate gently with PBS warmed at least to room temperature:
    Number of wells Volume
    96 0.2 mL
    48 0.5 mL
    24 1 mL
    12 2 mL
    6 3 mL
  3. Carefully remove PBS and add crystal violet solution. Incubate 10 minutes at room temperature:
    Number of wells Volume
    96 50 uL
    48 100 uL
    24 200 uL
    12 500 uL
    6 750 uL
  4. Wash plate 2x in tap water by immersion in a large beaker. Be careful not to lift off cells. Change tap water between washes.
  5. Drain upside down on paper towels, than add 1% SDS to solubilize the stain:
    Number of wells Volume
    96 100 uL
    48 300 uL
    24 600 uL
    12 1 mL
    6 1.5 mL
  6. Agitate plate on orbital shaker until color is uniform with no areas of dense coloration in bottom of wells.
  7. Read absorbence of each well at 570 nm.