Herskowitz Lab Protocol
From : JMG/RKT 7/96


Silver Stain

Make 25% glutaraldehyde [Kodak #8648]

  1. Dilute from stock
  2. Mix 600ml with 4 teaspoons activated Norite Ex charcoal
  3. Stir freq 5'
  4. Filter through large Buchner funnel, Whatman D1 11.0cm filter paper
  5. Repeat above three with 2 teaspoons charcoal & clean filters
  6. Refilter into brown bottle, store at 4°C

Protocol for 1 gel

  1. Gel into 200ml of 5% MeOH 10% HOAc for 1hr
  2. Wash several times with nanopure H2O; preferrably leave o/n; wash at least until destain smell gone
  3. 5% glutaraldehyde reduction for 30' (20ml of 25% made to 100ml)
  4. Rinse several times with nanopure H2O;until all traces gone
  5. Stain with for 10'

    21.0ml 0.36% NaOH [=1.8g/500ml]
    1.4ml NH4OH (58% is reagent ...)
    4.0ml 19.8% AgNO3 [=20g/100ml]
    --> to 200m

  6. rinse 2X nanopure H2O; let sit in third rinse 10'
  7. develop with :

    2ml 1.0% citric acid
    0.25ml formaldehyde (38%)
    to 500ml

Important Tips

  • Use glassware not plastic - especially the tray for the gel must be glass
  • Always wear gloves
  • Shake gently
  • Do excessive washes with water to remove fixatives
Reference : Wray et al. (1981) Anal Biochem 118:197-203.

Lab Home  |  People  |  Research Summary  |  Bibliography  |  Links  |  Protocols