Fix
3 * 5 min, 2* 45 min, or overnight
30% EtOH (300ml/L) with 10% HOAc (100ml/L)
1 __________ 2__________
Wash
3* 15 min, or 3* 5 min
30% EtOH (300ml/L)
1 __________ 2 _________ 3 _________
Rehydrate
1* 20 min dd Water
__________
Decrease Background
1* 2 min (for 1.5 mm gels: 4 min)
2.5 mM NaThiosulfate (0.62 g/L) (0.31g in 500ml)
Time: ____________
Wash
2* 5 min dd Water
1 _________ 2 __________
Expose to Silver
1* 30 min 0.2% AgNO3 (0.2 g/100 ml)
plus 0.8 ml 37% Formaldehyde per liter (80ul /100ml)
Time:__________
*** Dispose of in the silver waste container ***
Wash
1* 1 min dd Water
Revelation
2-10 min 3% Na2CO3 (3g/100 ml)
plus 0.5 ml formaldehyde 37% (50ul/100ml)
plus 20uM NaThiosulfate (800ul/100ml)
Time:
Stop
10 ml Acetic Acid
Try to anticipate how dark you want it and stop just before this point.
Wash dd Water