Mouse Heart Tissue Harvest for DNA Microarrays: Difference between revisions

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==Procedure==
==Procedure==
1. Anesthesize mouse with 0.02 ml 2.5% Avertin /gram body weight.
*1. Anesthesize mouse with 0.02 ml 2.5% Avertin /gram body weight.
2. Make incisions through the skin and the peritoneum and cut out approximately 100-200 mg of the tissues to be analyzed.
*2. Make incisions through the skin and the peritoneum and cut out approximately 100-200 mg of the tissues to be analyzed.
3. Immediately rinse the tissue in 1x PBS, blot dry, and weigh.
*3. Immediately rinse the tissue in 1x PBS, blot dry, and weigh.
4. Transfer the tissue into an eppendorf tube and freeze in liquid nitrogen.
*4. Transfer the tissue into an eppendorf tube and freeze in liquid nitrogen.
5. After all the samples are collected, they may be stored at -70°C or used for immunoprecipitation.
*5. After all the samples are collected, they may be stored at -70°C or used for immunoprecipitation.


2.5% Avertin (Chem Name = Tribromoethanol, Fluka #90710)
    Dilute 100% Avertin in warm PBS (55°C or higher) Filter sterilize, wrap in foil, store frozen aliqouts at -20°C Aliqouts good for ~3 weeks at 4°C


100% Avertin
'''2.5% Avertin''' (Chem Name = Tribromoethanol, Fluka #90710)
     Dissolve Avertin in tert-amyl alcohol (Sigma #A-1685) Wrap in foil, store at 4°C, good for years.  
    *Dilute 100% Avertin in warm PBS (55°C or higher)
    *Filter sterilize, wrap in foil, store frozen aliqouts at -20°C
    *Aliqouts good for ~3 weeks at 4°C
 
 
'''100% Avertin'''
     *Dissolve Avertin in tert-amyl alcohol (Sigma #A-1685)  
    *Wrap in foil, store at 4°C, good for years.
 
'''*Note:''' Tribromoethanol is extremely light sensitive. Its degradative products, HBr and dibromylacetaldehyde, are both very toxic.


*Note
    Tribromoethanol is extremely light sensitive. Its degradative products, HBr and dibromylacetaldehyde, are both very toxic.
==Notes==
==Notes==


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'''Relevant papers and books'''
'''Relevant papers and books'''


If this protocol has papers or books associated with it, list those references here. Below is an example for formatting purposes. See the [[OpenWetWare:Biblio]] page for more information.
No further references are available at this time
<biblio>
#Goldbeter-PNAS-1981 pmid=6947258
#Jacob-JMB-1961 pmid=13718526
#Ptashne-Genetic-Switch isbn=0879697164
</biblio>


==Contact==
==Contact==

Latest revision as of 14:08, 24 June 2009

Overview

List of reagents is not yet complete

Procedure

  • 1. Anesthesize mouse with 0.02 ml 2.5% Avertin /gram body weight.
  • 2. Make incisions through the skin and the peritoneum and cut out approximately 100-200 mg of the tissues to be analyzed.
  • 3. Immediately rinse the tissue in 1x PBS, blot dry, and weigh.
  • 4. Transfer the tissue into an eppendorf tube and freeze in liquid nitrogen.
  • 5. After all the samples are collected, they may be stored at -70°C or used for immunoprecipitation.


2.5% Avertin (Chem Name = Tribromoethanol, Fluka #90710) 

   *Dilute 100% Avertin in warm PBS (55°C or higher) 
   *Filter sterilize, wrap in foil, store frozen aliqouts at -20°C 
   *Aliqouts good for ~3 weeks at 4°C


100% Avertin 

   *Dissolve Avertin in tert-amyl alcohol (Sigma #A-1685) 
   *Wrap in foil, store at 4°C, good for years.

*Note: Tribromoethanol is extremely light sensitive. Its degradative products, HBr and dibromylacetaldehyde, are both very toxic.

Notes

No further notes are available at this time.

References

Relevant papers and books

No further references are available at this time

Contact

  • Who has experience with this protocol?

or instead, discuss this protocol.


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