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→Protocol
#Cut frozen tissue on a glass plate on dry ice. Place in a new round bottom eppendorf tube.
#Weigh frozen tissue samples, only need 20-50 mg of tissue. If there is too much cut it off and return the extra tissue to the -80. Record the weight of each tissue.
#Cool the centrifuge to 4C.#Add 20 uL/mg of RIPA (keep on ice) or other buffer to tissue (400-1000 uL)
#Add a stainless steel bead and keep tissues on ice. Homogenize using Qiagen Tissue Lyser (3 min at 25Hz for WAT/Liver and 5 min at 50 Hz for Muscle).
#Remove stainless steel bead or transfer the homogenized tissue to a new tube.
#Centrifuge at 14 000 RPM at 4C for 10 min
#Remove supernatant to clean tube. If lysing fat, try to avoid the floating fat cake. If necessary respin to clarify
