Good morning everyone
I have 2 students to be working on mice heart muscles. One student will be doing Western Blotting technique for her study (protein quantification). The second one will be working on CD36 cells.
After harvesting the hearts, what solution can they store the heart tissue in before use?
Thanking you in advance for your help.
For the western blot, I would just make lysate and store it at -80c until you are ready to use it. For the CD36 stain, do you plan to do IHC? Our lab usually just fixes heart tissue with 4% PFA for a few days before submitting for sectioning.
You can snap-freeze the tissues in liquid nitrogen and store at -80c until use or you can prepare tissue lysates and store them at -80c. For preparing and storing the lysates, we use a buffer containing 0.25M sucrose, 1mM EDTA, 20µM DTT (pH 7.2).
Just a note in addition to the suggestions of John and Shalinee. For immunoblotting, your lysis buffer should contain a protease cocktail and, if you intend to assess the phosphorylation state of any proteins (i.e., using anti-phospho antibodies) a phosphatase inhibitor cocktail. Keep all solutions cold and work on ice whenever possible. If you can, pulverize your heart tissue under liquid nitrogen using a mortar and pestle, transfer the powdered tissue to a hand homogenizer (e.g., Dounce or potter-elvehjem). Suspend and homogenize in lysis buffer. Centrifuge, aliquot supernatant, store aliquots frozen at -80.
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