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Membrane protein concentration

Membrane preparation cell culture 35S GTP assay CHO cells

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#1 Joon Kim

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Posted 07 February 2013 - 01:07 AM

I'm preparing membranes from CHO cells cultured in Corning 75cm2 flasks. My protocol for this is generic: Scrape cells at 90~100 confluence, homogenise and spin at 60,000g. My final pellet is resuspended in 3ml of buffer.

But when I got to my Bradford assay to determine protein concentration, I'm finding that from one flask, I get around 0.5mg/ml. Although I resuspend my pellet in 3ml, which might be high, I'm not entirely convinced that this is all I get out of it?

I was wondering if anyone has experience in this, and may have an idea of how much protein one may expect to get?

Thank you for your time.

#2 Curtis

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Posted 07 February 2013 - 10:40 PM

This doesn't look like a good protocol to me. usually fractionation protocols have more centrifugation steps. we have many protocols at the main protocol-online.org page. That's where I found my own protocol.





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