How to concentrate my small peptide - (Sep/26/2007 )
Hi All,
I use a yeast expression system to express a small peptide (~ 4kD), which secrets into the yeast culture medium (1 L). I will need to freeze-dry my peptide sample, but before do so I will have to bring down to total volume to 10-50ml and exchange the medium to pure water. I think of spin concentrators, but I did not find one with large capacity and equipped with <4 kD cutoff molecule weight membrane. Do you guys have any suggestions for my situation? Thanks a lot!
There are dialysis bags with low MW cutoff, the lowest is 500 Da- 2000 Da. (quiet expensive, ~$300). Dialysis vs DW or low salt solution then lyopholize.
SPE with C18 column or cartridge.
Thank you, guys. Genehunter-1, can the dialysis bags handle 1L of solution? K.B., does SPE stand for Solid Phase Extraction? I've never use a SPE. Could you please give a rough idea how much it costs and whether it can handle 1 L volume of solution? Thanks again, guys.
If you slowly freeze your sample and remove the ice on the top, this would seem to be the easiest way for you to initially concentrate your sample of large volume. I dont know if you can do that. But you want to prevent the formation of whole chunk of ice that freezes all the way.
Thank you, genehunter-1.
spectrum chemicals sells rolls of dialysis membrane with various mw cutoff. you can cut it to the size you need for your volume.
I don't remember exact prices. Box of 20 columns, each with 5 g of sorbent (20mL total column volume) should cost below 200 Euro. Of course you can buy even bigger columns or bulk media. If handled properly columns can be used up to 10 times with satisfactory results. If your peptide is in low abundance you can run large volumes of your solution through relatively small column with good yield.
Search BioForum for SPE and peptides - we've discussed this matter few times in last few months.