McClean:Western Blot

From OpenWetWare

(Difference between revisions)
Jump to: navigation, search
(Overview)
(Materials)
Line 3: Line 3:
==Materials==  
==Materials==  
-
* Item 1
+
* Beta- mercaptoethanol
-
* Item 2
+
* 4X LDS sample buffer(Invitrogen NP0008)
-
* Stock Solution 1
+
* Protease inhibitor(Roche catlog number: 11836170001)
-
* Stock Solution 2
+
* Phosphatase inhibitor(Fisher catlog number: 78420)
-
 
+
* 20X NuPAGE running buffer(Invitrogen NP0001)
 +
* Prestained Protein Ladder(Invitrogen catlog number:10748-010)
 +
* Magic Marker(Invitrogen catlog number:LC5602)
 +
* NuPage pre-cast gel
 +
* PVDF membrane(Invitrogen catlog number:LC2005)
 +
* Methanol
 +
* 20X NuPAGE buffer(Invitrogen NP0006)
 +
* 1M Tris, pH 8.0
 +
* 2.5M NaCl
 +
* Tween 20
 +
* Nonfat dry milk
 +
* Primary and secondary antibody
==Stock Solutions==
==Stock Solutions==

Revision as of 12:20, 30 May 2013

Contents

Overview

This is a protocol to perform rapid yeast protein prep for SDS PAGE and Western.

Materials

  • Beta- mercaptoethanol
  • 4X LDS sample buffer(Invitrogen NP0008)
  • Protease inhibitor(Roche catlog number: 11836170001)
  • Phosphatase inhibitor(Fisher catlog number: 78420)
  • 20X NuPAGE running buffer(Invitrogen NP0001)
  • Prestained Protein Ladder(Invitrogen catlog number:10748-010)
  • Magic Marker(Invitrogen catlog number:LC5602)
  • NuPage pre-cast gel
  • PVDF membrane(Invitrogen catlog number:LC2005)
  • Methanol
  • 20X NuPAGE buffer(Invitrogen NP0006)
  • 1M Tris, pH 8.0
  • 2.5M NaCl
  • Tween 20
  • Nonfat dry milk
  • Primary and secondary antibody

Stock Solutions

Stock Solution 1

  • This is a very simple solution, so we only need a one line description of how to make it.

Stock Solution 2

This is a more involved solution, so we will describe how to make it in several steps:

  1. Step 1
  2. Step 2
  3. Step 3

Protocol

  1. Step 1
  2. Step 2
  3. Step 3


Notes

Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!

  1. List troubleshooting tips here.
  2. You can also link to FAQs/tips provided by other sources such as the manufacturer or other websites.
  3. Anecdotal observations that might be of use to others can also be posted here.

Please sign your name to your note by adding '''*~~~~''': to the beginning of your tip.

References

Gietz, R.D. and R.A. Woods. (2002) TRANSFORMATION OF YEAST BY THE Liac/SS CARRIER DNA/PEG METHOD. Methods in Enzymology 350: 87-96.

Contact

or instead, discuss this protocol.

Personal tools