Removing fat from tissue preps
Paul S. Brookes.
brookes at uab.edu
Thu Jun 8 13:06:33 EST 2000
One thing you might consider is the speed of the spin. Microcentrifuges
are not much good for preparing anything for SDS PAGE. You can only pull
about 10-15,000 x g in a microcentrifuge at 14,000 rpm top speed. If you
use an ultracentrifuge you should be able to pull at least 60,000 x g,
which will make the fat layer much more pronounced and easier to pipet away.
Also if concentration is not a problem, try homogenising in a greater
volume of buffer to improve the separation. 100mg of tissue in 1ml buffer
is going to give you such a high protein concentration that the fat might
be sticking to your proteins, hence its inability to be removed. Is there
detergent in your buffer? This might also aid in removing, or at least
emulsifying, the fat.
Dr. Paul S. Brookes. (brookes at uab.edu)
UAB Department of Pathology, G004 Volker Hall
1670 University Blvd., Birmingham AL 35294 USA
Tel (001) 205 934 1915 Fax (001) 205 934 1775
The quality of e-mails can go down as well as up
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