EDC Impressum & DSE

Electrophoresis
Development & Consulting
Dr. Hanspeter Schickle
Wolfgang Gstrein

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SDS - Silver Staining Protocol (+native gels)

This staining recipe is not identical to the IEF-Silver staining!
IEF-silver-staining needs 20% (w/v) Trichloroacetic Acid (TCA) as a very first step!

This staining is for SDS- gels using the Laemmli-buffer system!
For EDC’s SDSGels please use our special staining protocol

Juli 13, 2018

step

reagent 

volume

time
(0.4-0.6 mm)

time
(1 mm)

1 Fixing

 40% (v/v) Ethanol / 10% (v/v) Acetic Acid
(160 ml EtOH / 40 ml HAc in 400 ml)
(place gel onto the solution in a glas tray, gel swims surface side down)

2 x 200 ml

2 x 20 min

2 x 30 min

2 Incubation

30% (v/v) Ethanol; 0.03% (w/v) Sodium Thiosulphate
10 mM Sodium Acetate; 0.125 % Glutardialdehyd (w/v)
(60 ml EtOH / 65 mg Thiosulfate / 300 mg NaAc / 1 ml Glutardialdehyde in 200ml)

200 ml

30 min

60 min

3 - 5 Washing

H2O dist (place gel on the glas tray bottom surface side up)

3 x 200 ml

3 x 20 min

3 x 30 min

6 Silvering

0.25 % (w/v) AgNO3 /0.03% (v/v) Formaldehyde
(500 mg AgNO3 (or 10 ml stock sol.) / 180 µl Formaldehyde in 200 ml)

200 ml

40 min

50 min

7,8 Washing

H2O dist

2 x 200 ml

2 x 1 min

2 x 10 min

10 Developing

2.5 % Na2CO3 (w/v) / 0.03% (v/v) Formaldehyde / 0.00075% (w/v) Sodium-Thiosulfate
(5 g NaCO3 / 180 ul Formaldehyde / 75 ul Sodium-Thiosulfate (2%, fresh!) in 200 ml)

2 x 200 ml

3 - 5 min

 10 - 20 min

11 Stopping
Preserving

10% (v/v) HAc, 10% (v/v) Glycerol
(20 ml HAc / 20 ml Glycerol in 200 ml)

200 ml

20 min

40 min

Drying

air dry , on the support-film
then roll on a polyester-sheet

---

16 h

3 h
then cover
with a plastic film

droptest

Droplet-test: within 3 seconds

Stock Solutions
1. 20% TCA: (Merck 1.00807.1000) Fill up this bottle with (about 850 ml) Bidest. This gives a “100%” solution.
    ---> 100 + 400 Bidest = 20%.
2. AgNO3: 5 % (20-fold) solution, 5 g in 100 ml.
3. Na2CO3: 25% (ten-fold) solution, 25 g (water-free!) in 100 ml.
AgNO3, Na2CO3 (water-free), Sodiumthiosulfate are from Merck
Ethanol, Acetic Acid, Formaldehyd (37%), Glutardialdehyde and Glycerol are from Merck

Abbr.: HAc = Acetic Acid, NaAc = Sodium Acetate