Knight:NuPAGE electrophoresis/Fast staining

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(Materials)
Current revision (12:24, 23 August 2007) (view source)
(Notes)
 
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==Procedure==
==Procedure==
#Add 100mL deionized water in staining tray.
#Add 100mL deionized water in staining tray.
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#Microwave staining tray loosely covered for 1 min on high.
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#Microwave staining tray loosely covered for 30 secs on high.
#Shake staining tray for 1 min on orbital shaker at room temperature.
#Shake staining tray for 1 min on orbital shaker at room temperature.
#Discard water.
#Discard water.
#Repeat steps 1-4 two more times.
#Repeat steps 1-4 two more times.
-
#Add 20mL SimplyBlue SafeStain.
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#Add 20mL SimplyBlue SafeStain (enough to just cover the gel).
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#*Need 100mL for staining tray.
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#Microwave staining tray loosely covered for 20 secs on high.
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#Microwave staining tray loosely covered for 45 secs on high.
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#*1 min seemed a bit too long.
#*1 min seemed a bit too long.
#Shake staining tray for 5 mins on orbital shaker at room temperature.
#Shake staining tray for 5 mins on orbital shaker at room temperature.
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#Gel can be stored in salt solution for several weeks.
#Gel can be stored in salt solution for several weeks.
-
This protocol seems to work pretty well but is not as sensitive as the [[Knight:NuPAGE electrophoresis/Slow staining]] approach.
+
==Notes==
 +
*This protocol seems to work pretty well but is not as sensitive as the [[Knight:NuPAGE electrophoresis/Slow staining]] approach.
 +
*To date, the best staining tray I've found is the lid of a 1000&mu;L pipette tip box.  Then use a piece of mesh that just fits inside the lide to either keep the gel in place while changing solutions or to move the gel to and from the light box.  This method requires smaller volumes of stain than the staining tray from Invitrogen.
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[[Category:Protein]] [[Category:In vitro]] [[Category:Protocol]]

Current revision

Contents

Overview

A fast protocol for visualizing bands on a polyacrylamide gel via coomassie-like staining.

Materials

Procedure

  1. Add 100mL deionized water in staining tray.
  2. Microwave staining tray loosely covered for 30 secs on high.
  3. Shake staining tray for 1 min on orbital shaker at room temperature.
  4. Discard water.
  5. Repeat steps 1-4 two more times.
  6. Add 20mL SimplyBlue SafeStain (enough to just cover the gel).
  7. Microwave staining tray loosely covered for 20 secs on high.
    • 1 min seemed a bit too long.
  8. Shake staining tray for 5 mins on orbital shaker at room temperature.
  9. Discard SimplyBlue SafeStain.
  10. Add 100mL deionized water.
  11. Add a kimwipe.
  12. Shake staining tray for 10 mins on orbital shaker at room temperature.
  13. Add 20mL 20% NaCl for at least 5 mins.
    • Only do this step if you aren't planning on drying the gel! Otherwise, when the gel dries, you will see a white salt precipitate.
  14. Gel can be stored in salt solution for several weeks.

Notes

  • This protocol seems to work pretty well but is not as sensitive as the Knight:NuPAGE electrophoresis/Slow staining approach.
  • To date, the best staining tray I've found is the lid of a 1000μL pipette tip box. Then use a piece of mesh that just fits inside the lide to either keep the gel in place while changing solutions or to move the gel to and from the light box. This method requires smaller volumes of stain than the staining tray from Invitrogen.
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