（交流）介绍6种针对免疫荧光染色的细胞固定方法

1、Methanol-Acetone Fixation
  Fix in cooled methanol, 10 min at –20 °C.
  Remove excess methanol.
  Permeabilize with cooled acetone for 1 minute at –20 °C.

2、Paraformaldehyde-Triton Fixation
  Fix in 3-4% paraformaldehyde for 10-20 min.
  Rinse briefly with PBS.
  Permeabilize with 0.5% Triton X-100 for 2-10 min.

3、Paraformaldehyde-Methanol Fixation
  Fix in 3-4% paraformaldehyde for 10-20 min.
  Rinse briefly with PBS.
  Permeabilize with cooled methanol for 5-10 min at –20 °C.

4、PEM-Ethanol Fixation
  Fix in PEM buffer for 10 min.
  Rinse twice, briefly, with PBS.
  Permeabilize with cooled ethanol for 5-10 min at –20 °C.
  PEM Buffer: 0.1M PIPES, 5 mM EGTA, 2mM MgCl2 .6H2O. Bring to pH 6.8, using NaOH solution .

5、Acetone - Triton X-100 Fixation
   Rinse cells 2 times with HBSS at 37°C.
   Fix cells by incubation in acetone at RT for 5 min.
   Rehydrate cells for 10 min in PBS.
   Permeabilize cells with in 0.2% Triton X-100 in PBS for 5 min.
   Rinse samples 3 times, for one minute each time, in PBS.

6、Methanol - Acetone Fixation
   Briefly rinse cells with PBS at 37°C.
   Fix cells with -20°C acetone-methanol(1:1) for 10 min (do it in -20°C freezer).
   Either:  a) airdry cells and store, or
b) Rehydrate cells in PBS for 10 min at RT.

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