Establish stable ionization spray conditions before you calibrate the MS. The intensity and stability of the ionization spray largely depend on the performance of the API source. Use the Plot Chromatogram tool to evaluate the ion source’s ionization spray stability.

IMPORTANT

  • Failure to maintain a stable spray may result in a poor calibration or diagnostic result and compromise the data quality.
  • If the spray becomes unstable with your analyte solution, return to this topic to evaluate the spray stability.

Prerequisites

  1. Set up a 500 µL syringe with the FlexMix calibration solution, if applicable.

Procedure

  1. From the Tune application, apply the following settings:
  2. Ion Source pane:
  3. - Current LC Flow (µL/min): 3
  4. Syringe pump settings:
  5. - Flow rate (µL/min): 3
  6. - Volume (µL): 500
  7. Ion polarity mode: Positive
  8. Data type: Profile
  9. (Optional) Set up the system to use an analyte solution. Check the following:
  10. Verify that the LC device or the syringe contains a sufficient amount of the analyte.
  11. In the Ion Source pane, verify the value in the Current LC Flow (µL/min) box.
  12. In the Define Scan pane, specify the scan type to evaluate the spray stability.
  13. Place the MS in On mode .
  14. The MS begins scanning and applies high voltage to the spray insert. A real-time mass spectrum appears in the Tune window.
  15. Turn on the flow for the calibration solution or analyte solution, as follows:
  16. For the calibration solution, select Syringe Off to turn on the syringe pump.
  17. The button name changes to Syringe On.
  18. For an analyte solution, turn on the flow from the LC device or the syringe pump.
  19. A real-time plot of the solution’s mass spectrum appears.
  20. Plot the full total ion current (TIC) and relative standard deviation (RSD) graphs as follows:
  21. Select the Plot Chromatogram icon to open the Plot Chromatogram dialog box.
  22. Figure Plot Chromatogram dialog box
  23. Select the Spray Stability checkbox to monitor the %RSD of the total ion current.
  24. Select the TIC option to plot the full TIC chromatogram. Then select OK to continue.
  25. The Plot Chromatogram tool generates a real-time graph (plot) of the full TIC where you can observe the signal stability and the effects of changes to various parameters. The tool also generates a real-time graph of the %RSD of the TIC for a 10 Da-selected ion monitoring (SIM) scan that is centered around the most abundant mass-to-charge ratio (m/z) in the current spectrum.
  26. Observe the %RSD graph and review the signal stability rating and maximum %RSD value.
  27. The following table lists the criteria for a stable spray in positive or negative ion polarity mode.

Spray Stability Requirements for the Calibration Solutions

Ion Polarity Mode

Acceptable Signal Stability Rating

Maximum %RSD (threshold)

Positive

Excellent or Good

15

Negative

Excellent or Good

15

NOTE

If the signal stability rating is poor or the %RSD value is above the threshold after attempting to optimize the API source parameters and positions, contact Thermo Fisher Scientific Technical Support for assistance.