Formaldehyde Fixation

Date Published: July 18, 2023

Formaldehyde fixation (for membrane associated components)

Fixative Preparation
  1. Dissolve 16 g paraformaldehyde in 80 ml dH2O by stirring at 70°C under fume hood.
  2. Add a few drops of 1N NaOH to depolymerize the paraformaldeyde.
  3. Adjust the pH to about 7.0. Cool down to room temperature and bring up to 100 ml with dH2O.
  4. Filter through a 0.45 µm Millipore filter and mix with an equal amount of double strength buffer.
  5. Divide into convenient aliquots and store frozen at -20°C. Discard unused thawed fixative.

Cell Fixation

  1. Rinse cells with PBS at room temperature.
  2. Fix in 3-4 % paraformaldehyde in PBS for 15 minute at room temperature.
  3. Wash 3-times 5 minute each with PBS containing 100 mM glycine.
  4. Permeabilize cells with 0.1% Triton® X-100 in PBS for 1 to 4 minutes.
  5. Rinse cells with PBS.

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