Protein binds to column.

Record: 758

Category: Column Problem
Product: KW
Sample:

Question
I am using a Shodex KW-803 for running protein samples on HPLC. Recently I found all my positively charged protein (at pH7.5) were sticking to the column. I want to wash them out but the pH range is 3-7.5. I’d like to know can I wash it with basic solution? If not, how can I clean the column?

Answer
What solvent are you using? Is it 100% water?
Cation polymers tend to be adsorbed on the silica-gel and thus they are often very difficult to be analyzed by KW columns.

What we could suggest at this point is to following:
Reverse the flow and introduce ~0.3M NaCl solution to clean the column, use 0.5 mL/min flow.
*do not leave NaCl solution in the column for a long time period since it may cause rust to the column housing.

As for analysis, conditions like below could be an alternative option.
http://shodex.net/index.php?seitenid=1&applic=1070

If you need to use KW column, other option could be to use [50mM NaCl + 15mM Dodecyltrimethylammonium chloride]. However, since Dodecyltrimethylammonium chloride is a cationic surfactant, this will permanently stay in the column and may change the analysis performance for other analysis.

 
 
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