Controlling the M5 MicroLC system with SCIEX OS software using contact closure
Contact closure can be used to control external devices that are not directly controlled by SCIEX OS software. A sample batch is first created in the SCIEX OS software for MS acquisition, and then a similar batch is created on an external LC device with the required methods and autosampler settings. The contact closure allows the start of MS acquisition to be triggered from the external device at the start of the LC gradient method. Here, use of contact closure to synchronize the M5 MicroLC system with a mass spectrometer controlled by SCIEX OS software is described.
Setting up the contact closure:
- Connect the contact closure cable from the back of the LC system (I/O - G1) to the AUX port of the mass spectrometer.
- First, click the Configuration tile on the SCIEX OS software home screen. Then, deactivate the current configuration. Select a mass spectrometer to use and click the Edit button (top right of screen). When the device dialog opens, click the Settings button. Select Contact Closure at the bottom.
- Next, click Test Device. A successful test will produce a message in green text. Click the Save button in the Device window and then Activate Devices by clicking the button at the top of the screen. On the status bar under devices, Contact Closure should now be present with a green check mark (see screenshot).
- Remember to turn off contact closure when you are switching over to a different LC system that is controlled within SCIEX OS software!
Setting up the MS system batch:
- Set up your batch in SCIEX OS software first. No LC method information needs to be entered in the LC Method column. Plate and vial location are also not required.
- When running your samples, you will see that contact closure is enabled in the right hand status bar.
- Make sure that the MS method duration is less than the LC gradient method runtime in the M5 MicroLC system method to maintain robust synchronization.
- Note that the LC information will not be automatically stored in the MS data file. Therefore, it is helpful to add custom columns to the SCIEX OS software batch so that the autosampler or LC methods used, or any other key information, can be saved in the data file.
Setting up the M5 MicroLC system method:
- The M5 MicroLC system will be controlled using Analyst® Software, so the software must be installed on the acquisition computer.
- Make sure SCIEX OS software services (ClearCore) are closed before starting the install process below.
- Install Analyst software 1.7.1.
- Install Eksigent Control Software (ECS) v4.3.
- Install Analyst Device Driver (ADD) v1.3.
- Install Virtual Terminal as needed.
- Configure PAL3 and ADD.
- Open the Eksigent Driver Configure Utility and activate the Eksigent System Driver. Click Apply.
- Now that the software is set up, launch Analyst Software and create a new hardware profile under the configuration section. Set up a hardware profile with an MS in simulation mode, and then add the LC components, the Analyst Device Driver application, and the two gradient pumps. Activate this hardware profile.
- Next, build an acquisition method. Because the MS is in simulation mode, you can select any scan type for the method. The source settings and other details do not matter.
- Add the gradient methods to be used in the appropriate places in the method.
- To add the autosampler (AS) method, first open the *.dam acquisition method in ADD, enter the desired settings for the method, and then close the method. The AS method should now show up in the MS acquisition method. Resave the method to store the full method.
- Make sure you select the right method type (direct inject or trap elute) when building the AS method.
- Make sure LC Sync is selected in the method for the Synchronization Mode.
- Set the duration in the MS method to less than that of the LC gradient. For simplicity, you can set the duration of this method very short (20 sec). However, if you want to save the pressure traces, they will be stored in the Analyst software data file. In this case, you want the MS duration to be long enough to capture the portion of the pressure trace you are interested in storing.
Setting up the M5 MicroLC system batch:
- Now a batch needs to be created in Analyst Software to run the M5 MicroLC system. Add the same number of samples to this batch in the same injection order as in the batch created in the SCIEX OS software.
- Select Build Acquisition Batch, then add your samples and select the method that contains the M5 MicroLC system method you want to run.
- Best practice is to copy the data file name and the sample name from the SCIEX OS software batch to the Analyst software batch to ensure you are keeping your two separate batches in sync.
- Submit to the Analyst software queue.
- Submit the MS batch to the queue in SCIEX OS software and then start the queue. The MS MicroLC system will display “Loading” status while waiting for the LC injection.
- Next, click Start in Analyst software to start the queue in Analyst and start the M5 MicroLC system. Always make sure the SCIEX OS software batch is started first, so it is waiting for the sync signals from the LC system to start.
Important timing considerations for different LC method types:
- Acquisition starts in SCIEX OS software when the M5 MicroLC system valve switches for Direct Inject. For Trap/Elute, acquisition will start either at the end of the Gradient 2 method, or at the M5 MicroLC system AS wash cycle, whichever comes first.
- When running the method in Analyst Software with the MS in simulation mode, acquisition in the Analyst software will start after the wash cycle of the M5 MicroLC system. This creates some timing considerations when building your methods to ensure correct results.
- You always want your SCIEX OS software method to be shorter than the LC method duration so SCIEX OS software is always waiting for the start signal from the LC.
- The Analyst software method should always be the same or shorter than the LC Method Duration minus the Wash Time.
Direct injection method
- Below is a typical timing diagram for the direct injection workflow.
Trap elute method
- Below is a typical timing diagram for trap elute workflow.
- It is very important to have the Wash cycle LONGER than the Gradient 2 method or the valve will switch too early. When running longer loading times with the Gradient 2, make sure you extend the washing time to ensure the Gradient 2 still completes before the Wash cycle is over. Alternatively run higher flow rates on Gradient 2 and shorten the run time, again to ensure it finishes before the Wash cycle ends.
Viewing the pressure traces after acquisition is complete:
- The Analyst software data file can be opened in SCIEX OS software to view the pressure trace. Open the file then select Show > UV/DAD/ADC channel and select the pump channel you wish to view.