Low molecular weight proteins below 30kDa - (Dec/11/2013 )

Hey everybody

I have a problem where I want to separate the LMW protein under 30 kDa. I have used Centriplus centrifugal filters (YM-30) with a molecular

weight cutoff (MWCO) of 30 kDa which were purchased from Millipore. However, when I try this out I am not able to separate LMW proteins. Everything stay on the top of the filter, no LMW below 30 kDa commes through. Have I bougth the right filters? In the paper: Electrophoresis 2004, 25, 1289–1298, An investigation into the human serum “interactome” are using same filter. I do not know why I am not able to separate the LMW below 30 kDa with these filters. Can you help me out? We believe this filter concentrate the sample but are not able to separate the LMW proteins.

Merci,

Kambiz

-Kambiz-

The cutoff is very approximate, and gradual, not sharp. What is your molecular weight? You may need a different filter.

-phage434-

I just want everything below 30kDa. If it is less it is not important as long it is not bigger MW. How ever nothing comes through. That is my main problem ...

-Kambiz-

Those filters are not really meant to separate proteins by size. As phage said, those MWCO numbers are very approximate and usually pretty conservative. I've concentrated protein mixtures containing free GFP (28kDa) in a 50 kDa MWCO filter and the GFP still does not go through even though it's not aggregated or associated with anything else.

If you really want to separate proteins by size, use gel filtration.

-labtastic-