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Star Republic: Guide for Biologists

Buffer exchange --- Desalting column

This protocol uses the PD-10 desalting column from Amersham biosciences.

1. Remove the bottom plug off new columns or the cap for re-used column.

2. Decant the buffer.

3. Equilibrate the column with 25 ml of the buffer you would like to exchange into. Add 5 ml of the buffer each time, let it drain, and repeat 5 times.

4. Add exactly 2.5 ml of the sample. Do not collect the first 2.5 ml flow through as the sample enters the column.

5. When the sample has completely entered the column, switch to 15 ml collection tubes at the bottom of the column.

6. Add exactly 3.5 ml of the buffer, collect the flow through, which contains the sample exchanged into the new buffer.

7. The column can be recycled: wash with 10 ml of dH2O, then 25 ml of 20% ethanol or PBS/0.1% sodium azide. Store in the wash buffer.