Silver Development protocol ( silver stain ) for 0.75mm polyacrylamide gels
Agitate gel twice with 50 % MeOH , 10 % acetic acid ( Fixer 1 ) for 15 min ea .
Rinse ( 1 min ) with 200 ml DW .
Agitate in 10 % EtOH , 5 % glacial acetic acid ( Fixer 2 ) for 6 min .
Agitate in distilled water .
Agitate twice ( 9 min each ) with 500 ml DW .
Agitate in 500 ml of 20 mg / L Na2S2O4 ( hydrosulfite [ dithionite ] , make fresh ) in distilled , deionized water ( DDW ) for 9min ( sensitization ) .
Pour off solution .
Agitate for 9 min .
Rinse 30 sec with DDW to remove excess AgNO3 .
Add 200 ml image developer ( 1ml 37 % formaldehyde per liter of 3 % sodium carbonate ) mixed with 200 ml 10g / L sodium thiosulfate .
Pour off developer .
Add 80 ml of stop solution ( 50 g tris , 25 ml glacial acetic per liter of distilled water ) .
Store gel in 10 % glycerol with bacteriostat ( 0.02 % NaN3 ) .
Without rinsing , add 200 ml , 0.1 % AgNO3 ( 200 mg ) in DDW .
Add 150 ml 37 % formaldehyde .
Agitate to desired staining intensity ( 3 to 6 min ) .
