Hi,
I have a silly question regarding buffer exchange columns (in that case the zebda desalting columns, from thermo scientific).
So as I understand the process is pretty straight forward :
1. open column, spin and discard storage solution.
2. Load, spin and discard the desired new buffer 4 times to equilibrate the columns.
3.Load your sample, spin and Harvest flow through.
Now here's what I don't get. Since we discarded the "new buffer" everytime during step 2, how can our sample be in the new buffer at the end of step 3? Shouldn't that just be your disalted original buffer?
When you listed the order of events, you answered your own question.
- "2. Load, spin and discard the desired new buffer 4 times to equilibrate the columns.
- 3.Load your sample, spin and Harvest flow through."
NO SAMPLE is loaded on the column until AFTER you fully equilibrate the media. Your Step 2 is the equilibration process. In Step 3, you load the sample onto the media (The sample is dissolved in the buffer), push it through the support, then collect it. Please review the steps again and it will be crystal clear to you.
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