In this video, we are going to teach you how to clean your HPLC column.
This is especially important to do if you want to add buffers or additives to your mobile phase. For instance, most people use ammonium acetate, formic acid, etc. And over time, if the column isn’t flushed out regularly, precipitates can form and plug your column up.
That's why we are going to show you 2 techniques to extend your column life as much as possible.
The first method is your daily maintenance method.
If you have a solenoid valve in your pump, you can select up to 4 solvents. A and B can be used as your mobile phase for regular operations. C and D can then be selected to be used as your solvents for cleaning. C and D would normally consist of water and some kind of organic solvent like methanol. You can simply do a gradient after your last run of 5% to 95% methanol at over an hour.
If you feel like your column needs a deeper cleaning, it would be best to do a reverse flush. The method for reverse flush would be to
- 95% water and 5% methanol for 10 column volumes
- Do a gradient of 5 to 95% methanol for 5 column volumes
- Hold 5% water and 95% for 10 column volumes
You want to run this at half your normal flow rates for your method.
The way that you know when to do deeper cleaning is to monitor your pressure build up on your column. Always note the backpressure on your method when you first start to use your column and if pressure kick up around 15-20% it’s probably a good idea to use the reverse flush.