Hello,
Does anyone have a protocol or perform serum or plasma collection from mouse bone marrow? There doesn't seem to be a well-described method in the literature - most I've seen rely on crushing/flushing bones and spinning the supernatant. Does anyone know if this also allows for collection of molecules that are matrix-associated, or does that require enzymatic preparation?
Thank you!
Bryce
Flushing the femur, followed by centrifuge @350g for 5-7 minutes will allow the cellular particle to settle down. Now you have plasma in supernatent.
For matrix associated proteins you may need enzymetic method.
For blood collection we do a cardiac puncture (you can find video on YouTube) on mice under anesthesia and collect whole blood in a 1.5 sterile Eppendorf tube. For serum, allow the blood to clot for 30 min. then centrifuge at 2,000xg for 10 min at 4C. Supernatant is the serum. Store in the freezer if not being analyzed immediately.
Plasma needs to be collected in a specific tube containing EDTA. Purchase the tube from the manufacturer (ask wherever is analyzing the sample what they prefer, some provide tubes, we purchased ours). Centrifuge tubes at 10,000xg for 5 min the day before use to ensure EDTA is down and will make contact with the blood (or samples will clot, this happened to us). Take some amount of the total blood that you collect from cardiac puncture and put it in the specific tube (I think ours has purple lid) and mix the blood so that the EDTA can prevent the clotting. The blood will start clotting quickly, so put the first blood you collect directly in the tube for the plasma and then the remaining blood in the sterile Eppendorf tube for clotting.
Not sure about bone marrow, other than I was shown how to retrieve it from the leg because this resulted in the greatest yield (maybe search this and see if that helps find a protocol?)
Flushing the femur, followed by centrifuge @350g for 5-7 minutes will allow the cellular particle to settle down. Now you have plasma in supernatent.
For matrix associated proteins you may need enzymetic method.
