The effective capture of the cell membrane protein is important for the sample preparation protocol and subsequent analysis by Western blot. When the comment on the pellet, and use a low volume of re-suspension buffer as possible to obtain a sample of the high concentration of protein.
Protein extraction procedure:
A. Start with 1 gram of tissue washed in cold PBS
B. Using liquid nitrogen to freeze the sample rapidly
c. Grinding tissue with a mortar
D. Place fabric now powder in 10 ml buffer decomposition
e. Using the Polytron, a mixture of soil and analyze the tissue buffer (explosion - 20 seconds)
f. Centrifuge for 15 minutes at 500 G at 4 degrees Celsius
g. Remove and save the swag
h. Homogeneity Pele again with further analyze 5ml buffer (20 seconds)
Me. Centrifuge for 15 minutes at 500 G at 4 degrees Celsius
j. Combining both supernatants
k. Centrifugation and toured in the 45 000 x G for 15 minutes
l. Wash precipitate twice in the buffer decomposition - Remove toured the wash
M. Resuspend in the re-suspension buffer
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