Procedure
The Solution from yesterday was run through the anion exchange column
- The column was cleared from ethanol by running 1L distilled water through it
- The column was stabilized with 1L pH10 CHES buffer
- The filtered solution was placed on the column
- 100mL of pH 10 CHES buffer were run after the protein
- Nothing came off and there was a dark band about 1in thick at the top of the column
- 200mL of 200mM NaCL in pH10 CHES buffer were run through the column
- Red band started to move and was collected in a erlenmeyer flask
- This solution was concentrated by running in the centrifuge for 30 minutes at 3000rpm and 4C
- The solution was placed in a centrifugal device with a membrane that lets salts and liquid through but not protein
- The concentrated solution was desalted by adding more pH 10 CHES buffer and placing it in the centrifuge again
- This step was repeated twice
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