Sodium Citrate Antigen Retrieval:
1. Place slides in a glass slide holder and fill in the rest of the
rack with blank slides (10 total) to ensure even heating.
2. Place rack in 600 ml of 10 mM sodium citrate, pH 6.0 in a
glass 2L-beaker. Mark a line at the top of the liquid on the
beaker.
3. Microwave for 20 min total, replacing evaporated water
every 5 min.
4. Cool slides for 20 min.
5. Wash 4 x 3 min in ddH2O, and 3 min in 1X PBS
Proteinase K Antigen Retrieval:
1. Make a fresh solution of:
25 ul of 20 mg/ml Proteinase K
2.5 ml of 1 M Tris-Cl, pH 8.0
0.5 ml of 0.5 M EDTA, pH 8.0
qs 50 ml with ddH2O
2. Incubate slides in a glass slide holder with solution at 37ºC
for 5 min (do not pre-warm Proteinase K solution).
3. Wash 3 x 5 min with 1X PBS.
Urea Antigen Retrieval:
1. Make a fresh solution of 1 M urea.
2. Place slides in a glass slide holder and fill in the rest of the
rack with blank slides (10 total) to ensure even heating.
3. Place rack in 600 ml of 1 M urea in a glass 2L-beaker. Mark
a line at the top of the liquid on the beaker.
4. Microwave for 10, 20 or 30 min total, replacing evaporated
water every 5 min.
5. Cool slides for 30 min to 1 hr.
6. Wash 4 x 3 min in ddH2O, and 3 min in 1X PBS.