Use of ÄKTA purifier for purification
Always get personal instructions before you use the ÄKTA purifier.
Before use, turn on the computer screen.
When the ÄKTA isn’t in use, all hose (A11, B1, B2 and A2) must contain 20% EtOH.
MonoQ/S
Wash hoses in buffer:
1) Put the A11 hose in buffer A (low salt) and the B1 hose in buffer B (high salt).
2) Start the UNICORN Manager on the computer.
3) Change to System Control in the bottom of the screen.
4) Choose Manual Pump, from the menu at the top.
5) Choose PumpWashPurifier in the list of instructions.
6) Attach InletA1 to A11 and activate B1.
7) Press enter or click Execute.
8) After a few minutes, flow rate will reset to 0.00 ml/min, but the system is still
in”Run”-position. Now press the End button in the top section of the screen.
Set up the system:
1) Choose loop size, depending on the sample volume, 5ml, 10ml or superloop.
5 and 10 ml loops are attached to INJECTION VALVE 1, between port 2 and
6. the superloop is washed and filled with water in both ends and is attached to
port 2 and 6. Top of the loop goes to port 6 and the bottom to port 2.
2) Place Eppendorf-tubes in the fraction-collector.
Equilibrate the column:
1) Change to System Controle in the bottom part of the screen.
2) Choose Manual Pump from the menu at the top.
3) Click Flow and enter 0.5 (ml/min). Press enter or click Execute.
4) Buffer A will now run through the column. Wait for stability in UV, pressure
and ion strength (about 5ml total). Stop the system manually by pressing End
in the top of the screen.
OR preferably create a Method to do the wash!
Load the sample:
1) The sample is loaded with a syringe, the same way as the loops.
Start the program:
1) Change to UNICORN Main Menu in the bottom of the screen
2) Right-click on the method in the correct folder and choose Run from the
menu. If you by accident double-click on the file, it will be opened in Method
Editor and it is necessary to go back to UNICORN Main Menu. If you cannot
find the file, you are probably in the wrong folder. Double-click on the yellow
folder with an arrow up, to get to the level higher, before you double-click on
the folder.
3) Follow the instructions on the screen as usual.
After ended program, wash all hoses in EtOH:
1) Add both A11 and B1 to 20% EtOH.
2) Change to System Control in the bottom of the screen.
3) Choose Manual Pump, from the menu at the top.
4) Choose PumpWashPurifier in the list of instructions.
5) Attach InletA1 to A11 and activate B1.
6) Press enter or click Execute.
7) After a few minutes, flow rate will reset to 0.00 ml/min, but the system is still
in ”Run”-position. Now press the End button in the top section of the screen.
Equilibrate the column back to 20% EtOH.
1) Change to System Controle in the bottom part of the screen.
2) Choose Manual Pump from the menu at the top.
3) Click Flow and enter 0.5 (ml/min). Press enter or click Execute.
4) 20% EtOH will now run through the column. Wait for stability in UV,
pressure and ion strength (about 5ml total). Stop the system manually by
pressing End in the top of the screen.
OR preferably create a Method to do the wash!
In the end: Turn of the computer screen by pressing the on/off button.
Superdex200
Start equilibrating the column the evening before you want to run it.
Wash the hose in buffer:
1) Put the A11 hose in buffer A, and B1 in ddH2O.
2) Change to System Control in the bottom of the screen.
3) Choose Manual Pump, from the menu at the top.
4) Choose PumpWashPurifier in the list of instructions.
5) Attach InletA1 to A11 and activate B1.
6) Press enter or click Execute.
7) After a few minutes, the flow rate will reset to 0.00 ml/min, but the system is
still in “Run”-position. Now press the End button in the top section of the
screen.
Equilibrate the column:
1) Change to UNICORN Main Menu in the bottom of the screen.
2) Start a pre-made Wash Program
3) Follow the instructions on the screen as usual.
Second Day:
Set up the system:
1) Attach a 250 or max a 500l loop to INJECTION VALVE V1between port 2
and 6.
2) Mount the black clamp for the Hamilton pipette.
3) Place Eppendorf-tubes in the fraction-collector.
Load the sample:
1) The sample is loaded with a Hamilton syringe.
Start the program:
1) Change to UNICORN Main Menu in the bottom of the screen
2) Right-click on the method in the correct folder and choose Run from the
menu. If you by accident double-click on the file, it will be opened in Method
Editor and it is necessary to go back to UNICORN Main Menu. If you cannot
find the file, you are probably in the wrong folder. Double-click on the yellow
folder with an arrow up, to get to the level higher, before you double-click on
the folder.
3) Follow the instructions on the screen as usual.
After ended program, wash hose again.
1) Add A11 to ddH2O and B1 to 20% EtOH.
2) Change to System Control in the bottom of the screen.
3) Choose Manual Pump, from the menu at the top.
4) Choose PumpWashPurifier in the list of instructions.
5) Attach InletA1 to A11 and activate B1.
6) Press enter or click Execute.
7) After a few minutes, flow rate will reset to 0.00 ml/min, but the system is still
in ”Run”-position. Now press the End button in the top section of the screen.
Equilibrate the column back to 20% EtOH.
1) Change to UNICORN Main Menu in the bottom of the screen.
2) Start a pre-made Wash program.
3) Follow the instructions on the screen as usual.
In the end: Turn of the computer screen by pressing the on/off button.
IF ANYTHING GOES WRONG:
Press the Pause button in the upper part of the screen and contact Bjørn or anybody
looking responsible in the vicinity.