Hello Gary,

One easy solution to reduce albumin burden is first grow the cells to confluence in regular 10% serum containing media. Then wash the cells once with blank media without serum and then substitute commercially available serum free media. Incubate the cells for a further period of 24-48 hours and collect the supernatant for mass spectrometric analysis. My understanding is even commercially available serum free media contain 1mg/ml albumin, which you can remove using blue sepharose or Caprylic acid precipitation. This is more practical than doing Caprylic acid precipitation directly on 10% serum containing media which has lot of albumin.

Hi Gary, I would recommend Blue Sepharose, it also binds BSA:

http://www.gelifesciences.co.jp/tech_support/manual/pdf/71708200ae.pdf

The kit from Pierce was very helpful for me in the past. See http://www.piercenet.com/product/albumin-depletion-kit

Beware though that some proteins and peptides may get stuck to the albumin and may get removed along the way. If this is the case for your molecule of interest, the best option is to use serum-free formulated culture media.

Thanks guys this is exactly what I was hoping for. I am hoping to generate two set of samples from cell culture media (i.e. media from low passage and from later passage cells) and then analyse it on Mass Spec. So as long as any "stuck" proteins are removed equally from each condition that should not be a problem (I am not looking for a particular protein in other words - just a "difference" in the Mass Spec results).

Thanks again!

Gary

For a different approach, if your cells tolerate it, you can also remove the FBS-containing media, wash the cells several times with PBS  to remove residual FBS, and add media that lacks the FBS  to the cells.  In most cases you can culture the cells this way for up to ~24 hours.

We have used Affi-Gel Blue from BioRad with good (but not perfect) results.  There was some leakage of dye from the column, which may or may not be a problem for you.  As others have pointed out, specificity is probably not absolute.

Forgot to mention:  You may want to read the article by Young et al. in Proteomics, 5, 4973-4979 (2005).  It includes a comparison of differen albumin purification methods.

Thanks Fritz, that's great info I will certainly check out the article.

Thanks again,

Gary

Try Caprylic acid precipitation. In mildly acidic conditions, the addition of short-chain fatty acids such as caprylic acid to serum will precipitate most serum proteins with the exception of the IgG molecules. See the link attached.

http://www.mechamlab.wustl.edu/Lab%20Web%20Page.data/Library/MethodsPDFs/AntibodyPurification%20.pdf

Sampath

It would be easier if you can use serum free media.

Thanks for the tip Sampath, in this instance I just need to remove the most abundant proteins (mainly BSA) but what you said is worth bearing in mind.

Hi Teodor-Doru it would indeed be easier if I could!

Thank you both,

Gary

So-called "serum-free" media for mammalian cells usually still contain some protein(s).  Molecules such as albumin are often needed as carriers for small molecules (e.g. lipids). 

some serum-free media are based only aminoacids and lipids, but not proteins.   

Hello Gary,

One easy solution to reduce albumin burden is first grow the cells to confluence in regular 10% serum containing media. Then wash the cells once with blank media without serum and then substitute commercially available serum free media. Incubate the cells for a further period of 24-48 hours and collect the supernatant for mass spectrometric analysis. My understanding is even commercially available serum free media contain 1mg/ml albumin, which you can remove using blue sepharose or Caprylic acid precipitation. This is more practical than doing Caprylic acid precipitation directly on 10% serum containing media which has lot of albumin.

Thank you Sampath that is very useful,

Thanks again,

Gary

Affi-Gel Blue, any simple affinity column to get out that huge BSA bulge-- maybe not complete, but certainly good enough to let you see bands without distortion!  We used for several publications of proteins exported or sloughed into media.

to follow this question I need to remove the BSA totally from culture supplemented with 2% BSA but without using serum free medium because my cells need it. 

Can anyone help me?