What are Autotune, Checktune, and Manual Tune and When to Run Them?

Answer

This Information Applies To: Agilent 6100 and 6400 Series LC/MS Systems

Issue 

Tuning a mass spectrometer involves optimizing the voltages applied to various optical elements in the ion path to achieve the required mass accuracy, resolution, and sensitivity that the user requires for their chosen application.

What are the differences between autotune, check tune, and manual tune and when is the best time to run them?

Background  

Tuning involves the use of a calibrant that consists of a set of perfluorinated compounds with known ion masses across a broad mass range (100-3000 m/z). Calibrant is infused into the MS during the tuning operation to optimize voltages for optimal transmission, expected m/z assignment, spectral peakwidth (m/z resolution), and detection using these known ion masses as a reference point.

For more details on calibrant/tuning mix, check out the following articles:

• Filling the Calibrant Bottle in 6200/6400/6500 Series LC/MS Systems
• Filling the Calibrant Bottle in Agilent 6100 Series Single Quadrupole LC/MSD Systems
• Checking the Calibrant Level in your Agilent LC/MS System
• Contamination of LC/MS Tune Mix - Diagnosis and Fixes
• Flushing the Calibrant System on your LC/MS System
• What Tuning Mix do I Use to Autotune my Agilent LC/MS System?

Resolution 

Autotune

• An automated tuning program that adjusts the MS for good performance over the entire mass range.
• See examples of the Autotune window in MassHunter Acquisition Version 10 (Figure 1), MassHunter Acquisition Version 12 (Figure 3), OpenLab CDS (Figure 5), and OpenLab ChemStation (Figure 7). 
• If the LC/QQQ autotune fails, check out this article What to do if my LC-QQQ Autotune Fails?

Checktune

• A program that checks the mass resolution and accuracy of the LC/MS. It provides feedback on which tune parameters require adjustment.
• Checktune does not tune the MS. The report indicates a pass/fail for different tune parameters. If any parameters are failing, end users would run Autotune.
• See examples of the Checktune window in MassHunter Acquisition Version 10 (Figure 1), MassHunter Acquisition Version 12 (Figure 3), OpenLab CDS (Figure 5), and OpenLab ChemStation (Figure 7).

Manual tune

• A program that allows users to tune the LC/MS by adjusting one parameter at a time until the desired performance is obtained. There are no reports generated, hence, no document trail is available. 
• See examples of the Manual Tune window in MassHunter Acquisition Version 10 (Figure 2) and OpenLab CDS (Figure 6), and the Advanced Override settings window in MassHunter Acquisition Version 12 (Figure 4).
• In MassHunter 12, Manual Tune is replaced by Advanced Override setpoints enabling some additional Tune parameters to be customized. Parameters (dependent on LC/TQ model) include Axis width, Axis gain and HED for Large Molecules (Figure 4).
  • Axis gain and Axis width -available on G6420A, G6460C, G6470A/B, G6495A/B/C
  • HED for Large molecule – available on G6420A, G6460C, G6470A/B, G6495A/B/C, G6475A

Where to perform tuning in different versions of software?

Select the software version that you are using from the list below for software-specific instructions.

MassHunter Workstation Data Acquisition Version 10 and older versions

MassHunter Workstation Data Acquisition version 12

OpenLab CDS Acquisition

OpenLab ChemStation Edition Online Session

MassHunter Workstation Data Acquisition Version 10 and older versions

Figure 1. Autotune and Checktune in MassHunter software
1. Tune context, 2. Autotune tab, 3. Autotune, 4. Checktune

Figure 2. Manual Tune in MassHunter software

MassHunter Workstation Data Acquisition version 12

Autotune

See Figure 3 for the location of each step in the Method Editor screen.

1. In the Method Editor window, click the TQ tab.
2. Click Tune > Autotune in the left pane.
3. Click Request tune control. This button locks control of the LC/TQ instrument. When clicked, tune setpoints are applied. If an action is not selected within the timeout period, the system will go back to the current method setpoints.
4. Select a polarity.
   1. If only Positive or Negative polarity is selected, then subsequent acquisition methods are only available with that polarity.
   2. If Both is selected, the system will automatically run in the positive mode followed by the negative mode.
5. Click Autotune the instrument. When the tune completes, review the report.
6. Click  Release tune control to release control of the LC/TQ instrument.
   
   

Checktune

1. Follow steps 1–4 in the Autotune section.
2. Click  Checktune the instrument. When the checktune completes, review the report.
3. Click  Release tune control to release control of the LC/TQ instrument.

Figure 3. Autotune/Checktune window in MassHunter Data Acquisition version 12
The numbers on figure 3 refer to steps listed in the Autotune section.

Advanced override settings (replaces old Manual Tune)

In MassHunter version 12, Manual Tune is replaced by Advanced override settings enabling some additional Tune parameters to be customized depending on the LC/TQ model being controlled.

1. In the Method Editor window, click the TQ tab.
2. Under the Method drop-down, select Advanced override settings.
3. Parameters to override from the current tune file can be checked depending on the model of LC/TQ being controlled, and an override value for that parameter can be specified.
4. When an Autotune is run the override values under the checked parameter will be used instead of the tune setting value.

Figure 4. Advanced Override setpoints window in MassHunter Data Acquisition version 12
1. TQ tab, 2. Advanced Override settings, 3. Parameter and input override value.

OpenLab CDS Acquisition 

Figure 5. Autotune and Checktune in OpenLab CDS software
1. Method layout, 2. Autotune tab, 3. Autotune, 4. Checktune

Figure 6. Manual Tune in OpenLab CDS software 

OpenLab ChemStation Edition Online Session

 
Figure 7. Autotune, Checktune and Manual Tune in OpenLab ChemStation software 
1. MSD Tune tab, 2.Tune drop-down menu, 3.Check Tune, 4. Autotune, 5. Manual Tune

When to Run Checktune vs Autotune 

Learn how to effectively operate your Agilent 6100 and 6400 Series LC/MS Systems:  LCMS-6400-1120sV2 - Tune and Calibrate Agilent 6400 Series Triple Quadrupole LC/MS with MassHunter Acquisition (Ver. 10.0) LCMS-6100-1101e - Tuning Agilent Single Quadrupole LC/MSD Systems in OpenLab CDS LCMS-6100-1102e - Tuning Agilent Single Quadrupole LC/MSD iQ System in OpenLab CDS eLearning courses available from Agilent education