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Lysis for WB with Ripa Buffer - too viscous, no pellet - urgent help needed, please

Lysis for WB with Ripa Buffer - too viscous, no pellet - urgent help needed, please - Western Blot Forum

Lysis for WB with Ripa Buffer - too viscous, no pellet - urgent help needed, please - Discuss western blotting and immunoblot in the western blot forum discussion board. Ask Questions about transfers, blocking, membrane antibody incubation and exposures.


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Old 02-05-2010, 08:08 AM
Pipette Filler
 
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Default Re: Lysis for WB with Ripa Buffer - too viscous, no pellet - urgent help needed, plea

In my lab we use a NP40, and then we add laemelli with DDT directly to the wells before we scrape. I am not getting consistent protein between my wells in my 96 well plates. I would like to use an alternative method of lysis so that I can get nuclear proteins, but I can quantify my proteins before using for a western. Can I use Ripa for this and then add my laemelli just before loading?
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Old 02-01-2013, 05:35 PM
Pipette Filler
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Default Re: Lysis for WB with Ripa Buffer - too viscous, no pellet - urgent help needed, plea

Thanks Laura, Just a quick question, do you recommend any voltage for sonication?
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Old 02-01-2013, 06:03 PM
Pipette Filler
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Default Re: Lysis for WB with Ripa Buffer - too viscous, no pellet - urgent help needed, plea

Thanks Laura, Just a quick question, do you recommend any voltage for sonication?
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Old 07-08-2013, 04:02 AM
Pipette Filler
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Default Re: Lysis for WB with Ripa Buffer - too viscous, no pellet - urgent help needed, plea

why dont we scrap plate in pbs and centrifuge them. then ad ripa and lyse. or add sample buffe and boil. i do this and never had prob.
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