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Following centrifugation, pellets were gently rinsed with 1 ml of the extraction buffer and centrifuged at 100,000 g for 20 minutes at RT.
Pellets were washed in homogenization buffer and centrifuged at 10200 g for 15 min to obtain a crude synaptosomal fraction.
After incubation for 15 mins with microbeads at 4 °C in the dark, cells were then washed with the buffer and centrifuged at 300 × g for 10 min.
The pellet was resuspended in an appropriate volume of isolation buffer, and centrifuged at 12,000×g for 8 min.
The resulting crude mitochondrial pellet was resuspended in 12% Percoll solution that was prepared in mitochondrial isolation buffer and centrifuged at 6900 g for 10 min.
The pellet was suspended in 1% Triton X-100 and 0.05-M phosphate buffer and centrifuged at 7800g for 10 min.
Tissue was homogenized in a lysis buffer and centrifuged at 15493 g for 20 minutes.
The lysate was added to 10 ml of buffer and centrifuged at 800× g for 15 minutes.
The harvested cells were boiled for 15 minutes in the sample buffer and centrifuged at 16,000 X g for 30 minutes.
The pellet fractions were washed in homogenizing buffer and centrifuged at 9200 × g for 15 min.
Thoracic aortas were homogenized in lysis buffer and centrifuged at 12000 g for 15 min.
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