Methanol Fixation for Immunofluorescence

Methanol fixation works by precipitating proteins, and as such it is a quick method (2-5)minutes is enough time for most antibodies/proteins). Diffuse proteins may be lost,however, in which case PFA fixation may be a better choice. We recommend that you try both types of fixation for your particular antibody/fusion protein when working out your initial immunofluorescence conditions.

MeS Buffer

100 mM MeS, pH 6.9

1 mM EGTA

1 mM MgCl2

Store at 4°C

Methanol Fix (100 ml)

10 ml MeS Buffer

90 ml Methanol

Store at –20°C and use on cells directly from freezer. Pour fix onto cells in dish and leave dish on the bench for 5 minutes, then wash repeatedly with PBS. There is no need for a permeabilization step following this fixation.