About the cell lysis methods - (Aug/27/2006 )

as i am going to lyse the cells and then do the SDS-page. At the beginning, i just want to folow the protocol, the procedures are:
1. Add lysis Buffer, then soncation

HOwever, my supervisor said that this method is not good, you should add SDS sample buffer into the sample and then boil for 10 minutes, he said it can lyse the cell as well

Is it better than the method showed in the protocol?
Thanks

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In my old lab, we used to boil for 10min and not sonicate as they never had a sonicator. This procedure worked for the westerns.

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really?
you mean SDS+boiling ==>work?

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yes, SDS, then boiling- works for westerns.

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but which one is better, thanks for yr suggestion

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i would say it depends on the protein you want to look at, and on the general purpose. some proteins dont like to bo boiled, and simply fall apart.. also, if you want to quickly measure the protein concentration of your protein sample, bme in your sample buffer may cause some trouble. otherwise, sds lysis+boiling works really well.

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I donot know which is better. U can try one of the methods and if there is problem then try out the other method. It may depend on the protein that one wishes to extract and what one wishes to use it for (like some assays).

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for most cells grown in culture the most useful method of lysis is traeting with detergents then physical shearing then freeze -thaw methods.
this is from my reading from (Antibodies A laboratory Manual by Ed Harlow and David Lane).

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I have always boiled my samples with sample buffer to prep them for western blots. I've never had any problems. Good luck!

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