SILVER STAIN OF PROTEIN GELS(Ⅱ)

1.Fix gel in 50% methanol for greater than 1 hour rinse gel in dH₂O 3 times 5 minutes each place back in 50% methanol for 1 hour.

2.Stain gel in 100 mls of stain solution for 30 minutes

3.Rinse gel in dH₂O 3 times 5 minutes each.

4.Place gel in Developer until bands are almost as dark as desired,sually 20' - 2 hours.

5.Stop developing and fix stain by placing gel in 50% methanol.Bands may continue to darken in methanol overnight,it's best to photograph gel immediately.

6.Photograph on light box.(for type 57 film use settings of 1/125 sec,f=22; for type 55 film use settings of 1/15 sec,f=11)

7.Wrap gel in prewet BioWrap® and allow to dry overnight

Stain solution

dissolve 0.8 g of AgNO₃ in 4 mls dH₂O

add a mixture of the following....

21 mls of 0.36% NaOH plus1.4 mls of concentrated NH₄OH (a little extra may be needed; add just enough to get silver to dissolve wait for lag!)then bring up to 100 mls with dH₂O (by adding 72.8 mls)

Developer

500 mls of dH₂O

2.5 mls of 1% Citric Acid (monohydrate!)

0.25 ml of 36% formaldehyde