Date post: | 27-Oct-2014 |
Category: |
Documents |
Upload: | nathanbeiber |
View: | 284 times |
Download: | 9 times |
ÄKTA™ and UNICORN™
A short introduction
2 /GE /
02/12/2007
UNICORN Basics
UNICORN is divided into four main sections:
UNICORN Manager: Used to organise data (result files, user set up etc) and configure the system for different optional components
Method Editor: Create and edit methods to control the instrument automatically
System Control: View run data and control the instrument manually (in real time)
Evaluation: Present the chromatographic data post acquisition in flexible, user defined formats
3 /GE /
02/12/2007
UNICORN Manager
Used to:• Create new users and define their level of access to the instrument. Once created, users will have their own folder where results and methods are automatically saved (highly recommended)
• Configure the software to recognise new hardware components
• Run Method Queues (methods run one after the other or to start at a certain time)
• Perform system diagnostics
4 /GE /
02/12/2007
UNICORN Manager Layout
Method filesMethod files Result filesResult files
Current folder where the
listed items are saved
Current folder where the
listed items are saved
5 /GE /
02/12/2007
Creating New Users
Select User Setup
from the Administration
Window
Select User Setup from the
Administration Window
6 /GE /
02/12/2007
Creating New UsersSelect New…Select New…
Enter User DetailsEnter User Details
Create a new folder where the methods and results for that user will be saved
Create a new folder where the methods and results for that user will be saved
7 /GE /
02/12/2007
Creating New UsersAfter the user has
been created, Select Edit…
After the user has been created, Select
Edit…
Change the Group to the desired level of
access to the instrument (see
below) *
Change the Group to the desired level of
access to the instrument (see
below) *
Select which folders will be shared with
other users and which System the user is allowed to operate
Select which folders will be shared with
other users and which System the user is allowed to operate
* An Administrator has full access to instrument including the creation of new users. To see the different levels of access select Access Groups
8 /GE /
02/12/2007
Handle files
Right-click on the file name!
Right-click on the file name!
Allows options such as copy, move, rename etc
9 /GE /
02/12/2007
System control workspace
CurvesCurves
Flow schemeFlow scheme
LogbookLogbook Manual instructions boxManual instructions box
Run DataRun Data
10 /GE /
02/12/2007
Manual instructions
The system is receiving instructions from UNICORN™. These instructions are either :
• automatically sent (by a method)
• manually sent (by the user)
Menu 'Manual'
11 /GE /
02/12/2007
Allows you to interact with the system manually, by giving appropriate instructions from a dialogue box, or interact with a running method.
Instructions are essentially the same as in Method Editor
System Control, Manual
12 /GE /
02/12/2007
Äkta Explorer 100 Air flowpath
13 /GE /
02/12/2007
Äkta Explorer 100 Air, main path
14 /GE /
02/12/2007
Flow direction valve
15 /GE /
02/12/2007
Injection valve, Load sample
16 /GE /
02/12/2007
Injection valve, Inject sample
17 /GE /
02/12/2007
Method Wizard
The Method Wizard is the easiest way to create chromatographic methods
The Wizard is interactive and flexible
Most of the parameters are defaulted to tried and tested values
The Wizard steps you through method creation in the same order as you would perform a run (technique selection, equilibration, sample injection, fraction collection, buffer gradient etc
18 /GE /
02/12/2007
Select The Method Wizard
19 /GE /
02/12/2007
Wizard Continued… Method and Column Selection Select the type of chromatography
Ion exchange, gel filtration etc
Select the column from the list (default
parameters such as flow rate and pressure
limits are set for that particular column)
Select the column position on the column
Valve (if applicable)
Flexible flow rates = flows that can be modified
at the beginning of each run
Flow Regulation = flows that will automatically
adjust downward if the column pressure
exceeds pre-set level
Buffer Prep = adding conc. buffer, salt, pH
modifier and water to four different buffer lines
allowing the system to mix the correct
proportion to each to attain a desired pH (if
applicable)
20 /GE /
02/12/2007
Wizard Continued… Select Wavelength and Buffers
Select the three UV wavelengths to monitor
during the run (UV-900 modules only)
Select the pump inlets for buffer A and buffer B
(only applicable to AKTA units with buffer
selection or buffer prep valves)
21 /GE /
02/12/2007
Wizard Continued… Equilibration
Select the starting concentration of buffer B
to equilibrate the column prior to injecting
the sample
Select the length of the equilibration (N.B. This
value is set as Column Volumes (CV) i.e if you
use a 1mL HiTrap column then it will
equilibrate for 5mL)
Instead of just using a Default value, you can
ask the system to ‘Watch Equilibration’. Select
the type of watch equilibration function from
the drop down menu (e.g. continue with the
method if the system reaches stable
conductivity)
A ‘Watch Function’ can be set on many different parameters – it
is simply defined as a conditional control that if a parameter is
met then the system proceeds to the next step
22 /GE /
02/12/2007
Wizard Continued… Sample Injection
Select the injection technique – Manual
injection is available for all systems. If you
have a P-960 Sample Pump, then other
options such as direct loading of the column
become available using the drop down menu
(next page of this handout).
Select how much buffer will be used to empty
(flush) the loop onto the column. Good practice
is to enter a value here twice as much as the
loop volume attached to the injection valve.
For example if you have a 1mL loop, enter a
value of 2mL here.
Remember to keep the syringe in place in the
injection valve until after the system has
injected the sample on the column (avoids
sample loss or air getting in the system)
23 /GE /
02/12/2007
Wizard Continued… Auto Sample Injection
If you have a Sample Pump (P-960), you can
select Sample Pump Direct Loading from this
drop down menu.
Select the volume of sample the sample pump
will inject on to the column (due to the dead
volume suggest values higher than 5mL).
Alternatively, if you have an Air Sensor on the
Sample Pump – tick the box ‘Inject All Sample’
– the system will continue to load until air is
detected
Tell the system which port your sample is
located (8-port valve designated line S1 to S8)
These two tick boxes tell the system to flush the sample line –
choose either to flush the lines to waste using (say) 20% ethanol
in sample line S8 or to flush the remainder of the sample onto
the column with 10mL of buffer A located in the S8 line
24 /GE /
02/12/2007
Wizard Continued… Unbound Protein Fraction Collection
If you have a Frac-950, you can select the
rack from this drop down menu (e.g. the
yellow rack = 18 & 30mm tubes).
Select the Wash Out Unbound Sample with 2
CV – continues to run equilibration buffer
(buffer A usually) to remove the unbound
protein from the column before the gradient
begins.
Tell the system to collect this unbound
fraction, i.e. what volume is to be collected in
each fraction and the tube type. If you select
30mm tube it will collect this fraction in 50mL
Falcon tubes.
The system remembers where the last fraction was collected
from the previous run (if the same method is used). Tell the
system if you want to continue collecting fractions in the next
available tube or recycle back to the First Tube.
25 /GE /
02/12/2007
Wizard Continued… Bound Protein Fraction Collection
Choose between Fixed Volume or Peak
Fractionation – Peak Fractionation only
collects when a peak is detected (user inputs
these parameters that define a peak on the
next step of the Wizard
Tell the system to collect this bound fraction,
i.e. what volume to be collected in each
fraction and the tube type. If you select 18mm
tube it will collect this fraction in 15mL Falcon
tubes in the centre of the yellow rack.
The system remembers where the last fraction was collected
from the previous run (if the same method is used). Tell the
system if you want to continue collecting fractions in the next
available tube or recycle back to the First Tube.
26 /GE /
02/12/2007
Wizard Continued… Gradient Elution Profile
Select the Elution Profile. Choose between
Linear Gradient, Isocratic or Segmented
(Step) (user inputs these parameters that
define a Segmented gradient on the next
step of the Wizard
If you select Linear gradient, tell the system
the final concentration in terms of % Buffer B
and how many CVs it will take to reach this
concentration
Select to Clean After Elution with 100% B (5CV) to remove any
strongly bound contaminant from the column. Typically do not
select Reequilibration as this is performed at the beginning of
the next chromatography run
27 /GE /
02/12/2007
Run Setup Mode
At the completion of the Method Wizard, the following page appears.
Notes and questions to give information to the users
Possibility to change all of the variables that were entered during the Method Wizard
28 /GE /
02/12/2007
Blocks = Chromatographic steps
Methods are divided into logical parts
Useful to import blocks from other methods
equilibrationequilibration Sampleapplication
Sampleapplication
Wash 1Wash 1 elutionelution Wash 2Wash 2 reequilibrationreequilibration
29 /GE /
02/12/2007
A group of instructions
Block equilibration
Main Method
Block equilibrationBlock wash
Block sample application
A method contains a main method and a set of blocks
Blocks
To use a block, it must be :
• created
• called
Main Method
Block equilibrationBlock wash
Block sample application
ExecuteBlock wash
30 /GE /
02/12/2007
Text Method ModeMethod: desaltscout
Flexibility using VariablesFlexibility using Variables
Adapt value during
Start Up Protocol
31 /GE /
02/12/2007
Text Method ModeMethod: desaltscout
Conditional Control using
Watch commands
IF a certain condition is met
THEN an action will follow
32 /GE /
02/12/2007
Start a method
Start from Main Menu,
• Select Methods,
• Select the appropriate method using right mouse button,
• Select Run
• Follow the Start Up Protocol and fill in as appropriate
Start from System Control Menu
• Select File
• Select Run
• Select appropriate method
• Follow the Start Up Protocol and fill in as appropriate
33 /GE /
02/12/2007
Open a Result in Evaluation
After the run, the result will be stored in Main Menu, Results
To open a Result from Main Menu
• Select the Result file with left mouse button and double click
To open a Result from Evaluation Menu
• Select File
• Select Open
• Select Result
• Select the appropriate Result file form the menu
34 /GE /
02/12/2007
HELP
On using ÄKTA and UNICORN
ASK
US