PD-10 Desalting Columns and LabMate Buffer Reservoir From GE Healthcare (formerly Amersham Biosciences)

The PD-10 Desalting Kit by GE Healthcare contains 30 desalting columns and 10 buffer reservoirs. The columns are pre-packed with Sephadex G-25 Medium which allows the separation of low (M_r < 1000) from high molecular weight compounds (M_r 5000). The box that the columns are shipped in can be converted to a stand which holds the columns, complete with an eluant-collecting tray. The reservoirs can be bought separately and used as convenient snap-ons to the multi-purpose syringe-styled columns. The columns are stable with a wide range of buffers since their working pH range is 2 – 13. The manufacturers also boast of typical protein yields of greater than 95% with less than 4% low molecular weight salts.

I regularly use these PD-10 desalting columns to purify crude Arabidopsis plant protein extracts. I extract my proteins in a strong urea – tris buffer amended with a number of protease and phosphatase inhibitors since I am interested in phosphorylation patterns and associated signaling. The PD-10 columns provide a convenient buffer exchange step where I can replace my inhibitor cocktail amended buffer with just the urea – tris buffer. This is useful for me since I don’t want the inhibitor cocktails to be present in critical protein digestion steps which follow my extractions. The buffer exchange is, however, done with a different amount of buffer so users should be careful to include this dilution factor (2.5/3.5) if they want to quantify their protein extract following desalting.

One thing I like about this product is the useful design where the box is conveniently converted to a column stand. This might seem trivial but previous experience with pre-packed desalting columns has been very frustrating since I’ve had to design my own stand to hold the columns. The buffer reservoirs are also very useful because they are re-usable and they allow equilibration of the column with larger amounts of buffer. This is actually very important to ensure efficient separation of proteins.

I’m not, however, very impressed with the high protein yields that the manufacturer claims. By measuring my proteins pre- and post-desalting, I find a significant loss in the total amount of protein. Crude protein extracts typically exhibit up to 5 mg/ml protein whereas the desalted aliquots of these same samples only have about 2 mg/ml. For my experiments, however, this is not so crucial since I don’t need anywhere near 2 mg/ml of protein to generate the amount of peptides I’m interested in after digestion. The only drawback is I run out of my desalted protein samples quicker!

The bottom line is for ease of use, buffer compatibility and efficient removal of salts this product is very good, but it could prove to be a let-down if your initial protein sample is not very much.