Storage of purified proteins

Curt Ashendel ashendel at
Wed Dec 14 15:11:18 EST 1994

On 13 Dec 1994 12:42:44 -0800, 
Gene Holowachuk  <geneh at USA.NET> wrote:

>Can anyone suggest some guidelines for the storage of highly purified
>recombinantly produced proteins. We have purified a couple of cytokines
>produced in E. coli, about 1 mg/l. What are the best ways for short term
>and long term storage - 50% glycerol, lyophilized, -70 oC, etc. We have
>found loss of activity when stored at 4 oC for longer than 1 week. Any
>suggestions will be appreciated.

If the intended purpose for your protein does not require activity, 
denature it in SDS or other denaturant and store at -20C or colder. This 
should prevent all proteolytic breakdown for long periods of time.

If the intended purpose for your protein requires activity, it should be 
stored as cold as possible (LN-2 is best, -70 to -100C next best, -20 for 
short term, 4C for a few hours at best); store as concentrated as possible; 
keep it from contacting skin and other sources proteases (i.e. use clean or 
better yet - sterile storage tubes); avoid repeated thawing and freezeing 
cycles.  The future use and stabilizing efficacy determines whether or not 
to include stabilizing agents such as 10 to 50% glycerol, non-ionic 
detergents, thiol reducing agents, and other proteins (e.g., BSA).  There 
is no general rule here, except that each of these help for some proteins 
and usually their stabilizing affects combine (additively or better).

If you can retain 100% of its activity as after lyophilization and 
rehydration, this would be the  best method.  Frequently this kills 
activity, however.  Avoid glycerol, most non-ionic detergents, and high 
salt concentrations in your protein solution before freeze-drying if you 
choose this method.

If you can store your protien as an suspension of precipitated 
protein in a high concentration of ammonium sulfate this will often work to 
stabilize many proteins to conformational and protease type degradation.  
Such suspensions often are stable for long times at 4C or -20C.  Some 
commercial preparations are shipped this way.  This works best for very 
concentrated preparations.

All solutions should be buffered to a pH freindly to your protein.

Personnaly, I like storing in small single use aliquots at -100C, usually 
in buffered 20% glycerol.  This works for most intracellular animal 

Curt Ashendel
Purdue University
West Lafayette, IN
ashendel at

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