[求救] microtubule sedimentation and cushion buffer
Why the sedimentation requires cushion ?
Could anybody tell me about the principle of cushion buffer for microtubule
sedimentation?
I has established an in vitro microtubule reassembly system from sperm
axoneme. Although a typical sigmoid curve of tubulin assembly could be
observed in turbidimetric assays, I still coud not recover reassembled
microtubules by ultracentrifugation. I didn't use cushion buffer to
sediment microtubules but the pellet is actually visible.
However, SDS-PAGE analysis is totally smear. Is cushion buffer essential?
Does ultracentrifugation crush reassembled microtubules permanently
so that I can't resolve reassembled microtubules in SDS-PAGE?
If cushion buffer is essential for sedimentation assays, how many volume
ratios and density shoud be adequate to pellet microtubules?
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