Tissue Protein Extraction Buffer (1X, 100 ml; Part No TEB-1)

Tissue Protein Extraction Buffer

For extraction, downstream immunoprecipitation and Western blotting of tissue proteins.  This buffer is non-denaturing, and maintains protein-protein complexes.  (1X HEPES-based, pH 7.5).  It is ELISA and immunoprecipitation compatible.   

Provided:  100 ml volume.  Sufficient for 20 ml of minced tissue.

Specifications:

• Extracts approximately 90% of proteins from tissues, including transmembrane proteins.
  
• Simple and rapid protocol for tissue protein extraction.  Use mini-homogenizer-pestle, microcentrifuge tubes and table top microcentrifuge.
  
• Maintains protein-protein complexes for downstream immunoprecipitation and pull-down experiments.  ELISA compatible.  Non-denaturing.
  
• Can be applied directly to extract tissue proteins for Western blotting.
  
• Compatible with Bradford assay.
  
• Absence of amines in the buffer allows for NHS-Ester conjugation and biotinylation.

1. Pulverize approximately 90 µL of tissue.  Place tissue in a 1.5 ml round bottom microcentrifuge tube.
   
2. Add general phosphatase and protease inhibitor cocktails to 500 µL of ice-cold Tissue Protein Extraction Buffer.
   
3. Add 500 µL Tissue Protein Extraction buffer with inhibitors to pulverized tissue.
   
4. Homogenize tissue with a mini pestle-homogenizer using 15 strokes, 3 seconds/stroke on ice.
   
5. Centrifuge 12000g for 15 min at 4^oC.
   
6. Remove supernatant (without lipid layer) and transfer into another 1.5 ml tube.
   
7. Centrifuge again at 12000g for 15 min at 4^oC.
   
8. Transfer supernatant to another tube.  The supernatant fraction contains the extracted proteins.
9. The Bradford assay can be used to quantitate extracted protein concentration.