Thermo Fisher Scientific 

TSQ Series - Installation Procedure P/N 70111-97117 Revision C 07.05.2012

 

FD Ayres 2 of 21 70111-97117 rev C

S/N TQU Service Order #

Service Engineer Start Date

Instrument Type: TSQ Series Vantage TSQ Series Ultra TSQ Quantum Access Max

The TSQ Series Installation Procedure is a working document that is provided to the customer at the time of instrument sign-off. Together with this document, the customer should receive the following documents as a stapled bundle: • Installation Service Report • Diagnostic test results for System Control • Diagnostic test results for Ion Source Board • Diagnostic test results for RF System • Diagnostic test results for Analyzer Control Board • Calibration report for positive ion mode • Calibration report for negative ion mode (if it is required) • Signal-to-noise report for ESI mode • Signal-to-noise report for APCI mode

TSQ Series Performance Specifications ESI Specification at 0.7 Da FWHM Resolution – [TSQ Quantum Access Max only] An on-column loop injection of the volume listed in Table 1 of a solution of 200 fg/µL reserpine into a 300 µL/min flow of (70:30) methanol (with no acid)/water (with 0.05% formic acid) will produce the minimum signal-to-noise ratio listed in Table 1 for the transition of the protonated molecular ion at m/z 609.3 to the fragment ion at m/z 195.1 when operated in the selected reaction monitoring mode (SRM). A minimum of 3 injections with a recommended variation of less than 15% in peak height must be demonstrated between 3 consecutive injections. The Q1 and the Q3 will have a resolution set to 0.7 Da FWHM. ESI Specification at 0.2 Da FWHM Resolution – An on-column loop injection of the volume listed in Table 1 of a solution of 200 fg/µL reserpine into a 300 µL/min flow of (70:30) methanol (with no acid)/water (with 0.05% formic acid) will produce the minimum signal-to-noise ratio listed in Table 1 for the transition of the protonated molecular ion at m/z 609.3 to the fragment ion at m/z 195.1 when operated in the selected reaction monitoring mode (SRM). A minimum of 3 injections with a recommended variation of less than 15% in peak height must be demonstrated between 3 consecutive injections. The Q1 will have a resolution set to 0.2 Da FWHM and the Q3 will have a resolution set to 0.7 Da FWHM. APCI Specification at 0.7 Da FWHM Resolution – [TSQ Quantum Access Max only] An on-column loop injection of the volume listed in Table 1 of a solution of 200 fg/µL reserpine into a 300 µL/min flow of (70:30) methanol (with no acid)/water (with 0.05% formic acid) will produce the minimum signal-to-noise ratio listed in Table 1 for the transition of the protonated molecular ion at m/z 609.3 to the fragment ion at m/z 195.1 when operated in the selected reaction monitoring mode (SRM). A minimum of 3 injections with a recommended variation of less than 15% in peak height must be demonstrated between 3 consecutive injections. The Q1 and the Q3 will have a resolution set to 0.7 Da FWHM. APCI Specification at 0.2 Da FWHM Resolution – An on-column loop injection of the volume listed in Table 1 of a solution of 200 fg/µL reserpine into a 300 µL/min flow of (70:30) methanol (with no acid)/water (with 0.05% formic acid) will produce the minimum signal-to-noise ratio listed in Table 1 for the transition of the protonated molecular ion at m/z 609.3 to the fragment ion at m/z 195.1 when operated in the selected reaction monitoring mode (SRM). A minimum of 3 injections with a recommended variation of less than 15% in peak height must be demonstrated between 3 consecutive injections. The Q1 will have a resolution set to 0.2 Da FWHM and the Q3 will have a resolution set to 0.7 Da FWHM.

Table 1. TSQ Series performance specification for ESI and APCI modes Instrument Sample Loop ESI S/N ESI 0.7 ESI 0.2 APCI S/N APCI 0.7 APCI 0.2TSQ Vantage Series 5 µL 6000:1 No Yes 1000:1 No Yes TSQ Quantum Ultra Series 5 µL 3000:1 No Yes 300:1 No Yes TSQ Quantum Access Max 5 µL 1500:1 Yes No 200:1 Yes No

FD Ayres 3 of 21 70111-97117 rev C

Instrument Setup 1. Use the lifting bars to set the instrument on the bench top.

2. Lower the two anti-skid feet to secure the instrument to the bench top. 3. Install the mechanical pump(s) and mist filter(s) with oil return lines. Fill the pump(s) to the appropriate level with

pump oil. Ensure that the blue exhaust hose is vented to a dedicated fume exhaust system. The exhaust system for the mechanical pump(s) must be able to accommodate a minimum flow rate of 1 L/min.

4. Install the IonMax source. Install the drain, the Drain Hose Adapter, and the 1-in. ID drain hose. Attach the

opposite end of the drain hose to the waste bottle. Ensure that the waste bottle is vented to a dedicated fume exhaust system. The exhaust system for the IonMax source must be able to accommodate a flow rate that does not exceed 30 L/min.

5. Use a separate waste container for the LC waste. Ensure that the waste container for the LC waste is vented to a dedicated fume exhaust system.

6. Connect the gas tubing for the argon and nitrogen gasses. The pressure for nitrogen and argon gas must be regulated to the value shown on the Foreline Cover Plate located next to the gas connectors. The Nitrogen to Vent connection is only to be used when the instrument is vented. Nitrogen to Vent should not be connected with tubing until the instrument is vented. It is not required to vent the instrument with nitrogen. Use this vent feature only if the customer requests that it be used.

Nitrogen for Vent 35 kPa Max (5 psi Max)

Argon In 135± 70 kPa (20 ± 10 psi)

Nitrogen In 690± 140 kPa (100 ± 20 psi)

FD Ayres 4 of 21 70111-97117 rev C

7. Connect the main power cable. 8. Install the PC and the Ethernet switch. Ensure the PC is set to the correct voltage. Turn on the PC.

9. The HESI-II probe is preinstalled with a 32-gauge metal needle. Confirm that the ESI needle protrudes 1.0 mm

from the tip of the nozzle and that the metal needle protrudes 0.5 mm from the end of the ESI needle and that the metal needle is not bent.

10. Install the HESI-II probe in the IonMax source. Verify that the 1/8-inch tubing for the Sheath and the Aux gas

are connected to the correct inlets located on the probe. The inlets are labeled A and S on the probe. 11. Turn the Main Power breaker switch to the On position. 12. Turn the Vacuum switch to the Operating Mode position (the TSQ Quantum Access Max does not have a

Vacuum switch). 13. Turn the Electronics switch to the Operating Mode position. The Instrument Manager will start the download

process. Confirm that the Instrument Manager displays the correct instrument type after communication is established.

Note. To troubleshoot communication problems refer to FAST 2005.026.

14. Confirm the following Power Entry Module LEDs are green: Forepump On, Vent Valve Closed, and Pass Ether link OK.

15. Confirm the Front Panel – Communication LED changes from flashing yellow to green. Pass

16. Confirm the Front Panel – Vacuum LED is green. The conditions for the Vacuum LED to turn green Pass are: • Fore pressure <2.5 Torr • Turbomolecular pump >90% of full rotational speed of 750 Hz • Ion gauge pressure is <7.0 × 10-4 Torr

17. Confirm the divert valve switches correctly. The Load and Inject LEDs should toggle between the Pass Load and Inject modes.

18. Confirm the syringe pump functions correctly. The Syringe Pump LED is green when the syringe Pass pump is running.

19. Browse to the C:\Thermo\Instruments\TSQ\system\programs folder and launch TSQ Tune enabled for Full Instrument Control. Do not add a shortcut to the Desktop. The customer is expected to use EZ Tune.

20. Set the Capillary Temperature to 270 ºC and allow the system to pump down. The instrument should reach a pressure of <3.0 × 10-6 Torr within 24 hours. The normal operating pressures for the instrument are: Ion transfer tube orifice sealed: Forepump pressure <0.05 Torr Ion gauge pressure <3.0 × 10-6 Torr

FD Ayres 5 of 21 70111-97117 rev C

Observed fore pressure (Torr)

Observed ion gauge pressure (Torr)

Note. To troubleshoot vacuum problems refer to FAST 2004.082.

21. Set the instrument to On. Confirm the Front Panel – System LED is green. The conditions for Pass the System LED to turn green are: • Vacuum LED is green • Instrument is in On mode • All interlocks are closed (readback 104 shows no fault ) • Pressure watchdog reads <6.0 × 10-6 Torr (readback 108 shows no fault ).

22. Run diagnostic tests in the following order. After each diagnostic test, save the results. The diagnostic test results are saved in the C:\Thermo\Instruments\TSQ\system\dgs folder.

Note. The Pressure Calibration diagnostic scripts are run in Manufacturing and do not need to be run in the Field. A regulator with a range between 0 and 100 psi is required to run the Pressure Calibration diagnostic scripts.

• System Control Pass • Ion Source Board Pass • RF System Pass • Analyzer Control Board Pass

23. Choose the Global Parameters tab and confirm that the following drift parameters are set to the correct values: Q1 Mass Correction (AMU/Degree) 0.008 Set Q3 Mass Correction (AMU/Degree) 0.012 Set Q1 Resolution Correction (AMU/Degree) -0.008 Set Q3 Resolution Correction (AMU/Degree) -0.010 Set

If the parameters are set to zero, use the Service Utility Tools to set the correct values (refer to FAST 2001.064).

Auto Tune and Calibrate The Auto Tune and Calibrate procedure is run with a low flow rate infusion of calibrant using the syringe pump. 1. Set-up the instrument for infusion of 1,3,6-polytyrosine via the syringe pump.

2. Remove the spray shield. 3. Set the instrument to Profile mode. 4. Set the Ion Source parameters to values specific to run Auto Tune and Calibrate:

Spray Voltage 3000 to 4000 Vaporizer Temperature 0 Sheath Gas Pressure 0 Aux Gas Pressure 0 Capillary Temperature 270

5. Set the Multiplier Gain to 300,000. 6. In the Define Scan view set parameter values so that they are the same as in the following figure.

FD Ayres 6 of 21 70111-97117 rev C

7. Use the syringe pump to infuse 1,3,6-polytyrosine at a flow rate of 1 to 3 µL /min. 8. Adjust the position of the HESI-II probe until a stable spray and a stable ion beam are obtained. Expect a

signal intensity of >1 × 107 counts. Confirm that the spray is stable by running the System Evaluation – API Stability Test. The results must be excellent. If the intensity of the polytyrosine is too high (saturated peaks with an intensity >2.6 × 108 counts), dilute the polytyrosine with methanol.

9. Run Auto Tune & Calib for both Q1 and Q3. 10. If the customer does not intend to operate the instrument in negative ion mode, copy the tune table to negative

ion mode. Then, go to step 11. If the customer will operate the instrument in negative ion mode, do not copy the tune table to negative ion

mode. Complete steps 11 to 14. Then, go to the section Negative Ion Mode Calibration. 11. Click Save Report to save the tune and calibration results. Examine the results and confirm that they are

normal. 12. Click Save Calib to automatically save the calibration file as the Current Calibration. You will then be

prompted to save the tune file. Save the tune file as ESI Polytyrosine Tune.TSQTune. 13. Open the Diagnostics Workspace. Then, choose File | Save Calibration As Default Factory. This file will

become the new default Factory Calibration file. 14. Run Gain and confirm the signal intensity is >1 × 107 counts.

Observed signal intensity (counts)

Negative Ion Mode Calibration (For customers that will operate in negative ion mode) 1. Set the instrument to negative ion mode. 2. Set the Ion Source parameters to values specific to negative ion mode calibration:

Spray Voltage 2500 or optimize automatically Vaporizer Temperature 40 Sheath Gas Pressure 40 to 80 Aux Gas Pressure 15 or optimize automatically Capillary Temperature 270

3. Infuse 1,3,6-polytyrosine at a flow rate of 20 to 30 µL/min. Expect a signal intensity of >106 counts. 4. Run Auto Tune & Calib for both Q1 and Q3. 5. Do not copy the tune table to positive ion mode. 6. Click Save Report to save the tune and calibration results. Examine the results and confirm that they are

normal. 7. Click Save Calib to automatically save the calibration file as the Current Calibration. You will then be prompted

to save the tune file. Save the tune file as ESI Polytyrosine Tune.TSQTune. 8. Open the Diagnostics Workspace. Then, choose File | Save Calibration As Default Factory.

FD Ayres 7 of 21 70111-97117 rev C

Optimize Peak Shape and Resolution (Required only for AM and EMR instruments) Note. Optimization of peak shape will result is a loss of signal intensity. Manual optimization should not cause a loss of signal intensity that is greater than 10%. 1. Set the instrument to Profile mode. 2. Set the Ion Source parameters to values specific for the optimization of peak shape and resolution:

Spray Voltage 3000 to 4000 Vaporizer Temperature 0 Sheath Gas Pressure 0 Aux Gas Pressure 0 Capillary Temperature 270

3. Set the Multiplier Gain to 300,000. 4. In the Define Scan view set parameter values so that they are the same as in the following figure.

5. Infuse 1,3,6-polytyrosine at a flow rate of 1 to 3 µL /min.

6. Select the Q1 page of the Device view and click Q1 Resolution to open the tune table. 7. Select the Auto Peak Width Setting check box. 8. Select the Peak Width Read Back check box. The peak width is displayed in the spectrum view as shown in the

following figure.

FD Ayres 8 of 21 70111-97117 rev C

9. In the Define Scan view with the Center Mass set to 182.082 and the Q1 Peak Width set to 0.7 observe the

intensity and peak shape at m/z 182.082. Make small incremental or decremental adjustments to the following parameter values only if it is necessary. Adjust the parameter values in the order they are listed starting with step 9a. Adjust the next parameter value only if it is necessary: a. Adjust the Spray Voltage and check if the peak shape improves. If the peak shape does not improve go to

step 9b. Otherwise, go to step 10. b. Adjust the Q1 DC Offset.

Note. It is usually not necessary to make manual adjustments to the resolution. 10. Click Q1 Resolution to open the tune table. Check if the peak width read back displays a value that is close to

0.7. If the read back is not close to 0.7, adjust the Value parameter spin box until the read back is close to 0.7. Hit the <Enter> key to apply the change.

11. Repeat steps 8 to 9 for the peak width set to 0.2. Hit the <Enter> key to apply the change. 12. Repeat steps 8 to 9 for the peak width set to 0.1. Hit the <Enter> key to apply the change. 13. In the Define Scan view set the Center Mass to 508.208. With the Q1 Peak Width set to 0.7 optimize both peak

shape and resolution. Hit the <Enter> key to apply the change. 14. Repeat steps 8 to 9 for the peak width set to 0.2. Hit the <Enter> key to apply the change. 15. Repeat steps 8 to 9 for the peak width set to 0.1. Hit the <Enter> key to apply the change. 16. In the Define Scan view set the Center Mass to 997.398. With the Q1 Peak Width set to 0.7 optimize both peak shape and resolution. Hit the <Enter> key to apply the change. 17. Repeat steps 8 to 9 for the peak width set to 0.2. Hit the <Enter> key to apply the change. 18. Repeat steps 8 to 9 for the peak width set to 0.1. Hit the <Enter> key to apply the change. 19. In the Define Scan view set the Scan Mode to Q3MS.

20. Select the Q3 page of the Device view.

FD Ayres 9 of 21 70111-97117 rev C

21. Optimize the peak shape and resolution for Center Mass 182.082 at peak widths 0.7, 0.2, and 0.1. Hit the <Enter> key to apply the change.

22. Optimize the peak shape and resolution for Center Mass 508.208 at peak widths 0.7, 0.2, and 0.1. Hit the <Enter> key to apply the change.

23. Optimize the peak shape and resolution for Center Mass 997.398 at peak widths 0.7, 0.2, and 0.1. Hit the <Enter> key to apply the change.

24. Click Save Calib to automatically save the calibration file as the Current Calibration. You will then be prompted to save the tune file. Save the tune file as ESI Polytyrosine Tune.TSQTune.

25. Open the Diagnostics Workspace. Then, choose File | Save Calibration As Default Factory.

Bake Out the HESI-II Probe, IonMax Source Housing, and Front-End Optics 1. Install the Hypersil GOLD aQ column at the ZDV union located on the HESI-II probe. 2. Confirm that the ion transfer tube is open. 3. Set the instrument to Profile mode. 4. Set the Multiplier Gain to 300,000. 5. Turn on the LC and set the flow rate to 300 µL/min for a flush mixture of (90:10) methanol (with no acid)/water

(with 0.05% formic acid). 6. Set the Ion Source parameters to values specific to bake out:

Spray Voltage 3250 Vaporizer Temperature 500 Sheath Gas Pressure 50 Aux Gas Pressure 20

Capillary Temperature 350 7. In the Define Scan view set parameter values so that they are the same as in the following figure. The

instrument is set to scan at a mass where no ions are expected in order to preserve the lifetime of the electron multiplier.

8. Bake out the HESI-II probe, IonMax Source Housing, and the front end optics overnight.

Demonstrate ESI Performance Performance is demonstrated by loop injection into an LC flow. 1. Set-up the LC and MS for manual loop injections.

FD Ayres 10 of 21 70111-97117 rev C

2. Use solvents of the appropriate purity (as specified in the TSQ Series Preinstallation Requirements Guide) to make the reserpine specification solution as described in the section Recipes.

3. Set the instrument to Centroid mode. 4. Turn on the LC and set the flow rate to 300 µL/min for the mixture of (70:30) methanol (with no acid)/water

(with 0.05% formic acid). 5. Set the Y probe position (up and down) between C and D. 6. Set the Z probe position (forward and backward) forward. 7. Set the Ion Source parameters to values specific to demonstrate ESI performance:

Spray Voltage 3600 Vaporizer Temperature 520 Sheath Gas Pressure 30 Aux Gas Pressure 10 (should not be adjusted) Capillary Temperature 340

8. In the Define Scan view set parameter values so that they are the same as in the following figure.

9. Confirm the Multiplier Gain is set to 2,000,000.

10. Click on the Display TIC button to open the Total Ion Current Plot. 11. Verify that the noise counts are: TSQ Vantage series <10 counts, TSQ Quantum Access Max <30,

TSQ Quantum series <50. 12. In the Diagnostics Workspace, run the Performance Specification – Enable diagnostic script. 13. Perform a loop injection of the reserpine specification solution and check the signal intensity by moving the

cursor over the top of the peak in the Total Ion Current Plot. The signal intensity is displayed at the bottom left of the window as shown in the following figure. Although there is no specification for signal intensity at installation, the instrument should achieve the intensity listed in Table 2.

The S-Lens parameter is displayed only when the software controls a TSQ Vantage series instrument

FD Ayres 11 of 21 70111-97117 rev C

Table 2. Expected ESI signal intensity for TSQ Series instrument at installation Instrument ESI signal intensity TSQ Vantage Series 1200 counts at 0.2 TSQ Quantum Ultra Series 8000 counts at 0.2 TSQ Quantum Access Max 4000 counts at 0.7

14. Monitor pump pressure. Pressure fluctuations should not exceed 5 bar. 15. If the loop injection does not achieve the minimum signal intensity, make small incremental or decremental

adjustments to the following parameter values. Adjust the parameter values in the order they are listed starting with step 15a and progressing to the next step only if it is necessary: a. Adjust the Y position (up and down) of the probe, then perform loop injections to check if the signal intensity

improves. If the signal intensity does not improve go to step 15b. Otherwise go to step 16. b. Adjust the Z position (forward and backward) of the probe, then perform loop injections to check if the signal

intensity improves. If the signal intensity does not improve go to step 15c. Otherwise go to step 16. c. Adjust the Sheath Gas Pressure. Changes to the sheath gas flow rate will affect the Vaporizer

Temperature. Wait 5 minutes after changing the Sheath Gas Pressure to ensure that the Vaporizer temperature has stabilized, then perform loop injections to check the signal intensity.

16. Click on the Acquire Data button to open the Acquire Data dialog box. Enter the File Name: ESI Performance Specification. Set the Acquire Time to 4 minutes.

17. Perform one loop injection of the reserpine specification solution every minute for a total of three injections. 18. Click on the Signal-to-Noise Test icon located on the desktop and enter the instrument serial number at the

prompt. 19. Select the Field radio button. 20. Select Open Raw File and open the ESI Performance Specification.RAW file. The signal-to-noise calculator will

automatically process the data and report the calculated signal-to-noise.

FD Ayres 12 of 21 70111-97117 rev C

21. Save the signal-to-noise result as a .PDF or .XPS file.

22. If the signal-to-noise ratio is less than expected, click on the Display TIC button to open the Total Ion Current Plot then, make the following adjustments to the probe position to reduce the signal intensity of the baseline noise: • Adjust the Y (up and down) position of the probe so that the probe is moved away from the ion transfer tube

orifice • If it is necessary, adjust the Z (forward and backward) position of the probe so that the probe is moved away

from the ion transfer tube orifice Then, go to step 16.

23. In the Diagnostics Workspace, run the Performance Specification – Disable diagnostic script.

Calculated ESI signal-to-noise

24. If APCI performance will not be demonstrated, then remove Performance Specification.dgs from the PC. If the instrument has the APCI option, go to the section Demonstrate APCI Performance. If the instrument has the extended mass range option, go to the section Installation Procedure for EMR Instruments. If the instrument has the accurate mass option, go to the section Installation Procedure for AM Instruments.

FD Ayres 13 of 21 70111-97117 rev C

Demonstrate APCI Performance 1. Install the APCI probe in the IonMax source. Ensure that the 1/8-inch tubing for the Sheath and the Aux gas

are connected to the correct inlets located on the APCI probe. The inlets are labeled A and S. 2. Install the Hypersil Gold aQ Javelin HPLC column to the inlet of the APCI probe. 3. Set the instrument to Centroid mode. 4. Turn on the LC and set the flow rate 300 µL/min for the mixture of (70:30) methanol (with no acid)/water

(with 0.05%formic acid). 5. Set the Ion Source parameters to values specific to demonstrate APCI performance:

Discharge Current 4 Vaporizer Temperature 450 Sheath Gas Pressure 30 Aux Gas Pressure 10 Capillary Temperature 320

6. In the Define Scan view set parameter values so that they are the same as in the following figure.

7. Confirm the Multiplier Gain is set to 2,000,000.

8. Click on the Display TIC button to open the Total Ion Current Plot. 9. In the Diagnostics Workspace, run the Performance Specification – Enable diagnostic script. 10. Perform a loop injection of the reserpine specification solution and check the signal intensity by moving the

cursor to the top of the peak in the Total Ion Current Plot. Although there is no specification for signal intensity at installation, the instrument should achieve the intensity listed in Table 3.

Table 3. Expected APCI signal intensity for TSQ Series instrument at installation Instrument APCI signal intensity TSQ Vantage Series 200 counts at 0.2 TSQ Quantum Ultra Series 1250 counts at 0.2 TSQ Quantum Access Max 1500 counts at 0.7

11. If the loop injection does not achieve the minimum signal intensity, make small incremental or decremental adjustments to the following parameter values. Adjust the parameter values in the order they are listed starting with step 11a and progressing to the next step only if it is necessary: a. Adjust the Y position (up and down) of the probe, then perform loop injections to check if the signal intensity

improves. If the signal intensity does not improve go to step 11b. Otherwise go to step 12. b. Adjust the Z position (forward and backward) of the probe, then perform loop injections to check if the signal

intensity improves. If the signal intensity does not improve go to step 11c. Otherwise go to step 12. c. Adjust the Sheath Gas Pressure, then perform loop injections to check the signal intensity.

12. Click on the Acquire Data button to open the Acquire Data dialog box. Enter the File Name: APCI Performance Specification. Set the Acquire Time to 4 minutes.

13. Perform one loop injection of the reserpine specification solution every minute for a total of three injections. 14. Click on the Signal-to-Noise Test icon on the desktop and enter the instrument serial number at the prompt. 15. Select the Field radio button.

The S-Lens parameter is displayed only when the software controls a TSQ Vantage series instrument

FD Ayres 14 of 21 70111-97117 rev C

16. Open the APCI Performance Specification.RAW file. The signal-to-noise calculator will process the data and report the calculated signal-to-noise.

17. Save the signal-to-noise result as a .PDF or .XPS file.

18. If the signal-to-noise ratio is less than expected, click on the Display TIC button to open the Total Ion Current Plot then, make the following adjustments to the probe position to reduce the signal intensity of the baseline noise: • Adjust the Y up and down) position of the probe so that the probe is moved away from the ion transfer tube

orifice • If it is necessary, adjust the Z (forward and backward) position of the probe so that the probe is moved

away from the ion transfer tube orifice. Then, go to step 12.

18. In the Diagnostics Workspace, run the Performance Specification – Disable diagnostic script.

Calculated APCI signal-to-noise

19. Remove Performance Specification.dgs from the C:\Thermo\Instruments\TSQ\system\dgs folder.

If the instrument has the extended mass range option, go to the section Installation Procedure for EMR Instruments. If the instrument has the accurate mass option, go to the section Installation Procedure for AM Instruments.

FD Ayres 15 of 21 70111-97117 rev C

Installation Procedure for EMR Instruments The TSQ Vantage EMR is calibrated for higher masses by appending the calibration file for polytyrosine. The procedure involves optimizing the lens parameters for the high mass peaks, performing a mass calibration, and then saving the calibration file. The procedure uses Ultramark as the calibrant.

Calibrate for EMR 1. Set the instrument to Profile mode. 2. Set the Ion Source parameters to the following values:

Spray Voltage 4000 Sheath Gas Pressure 0 to 3 Aux Gas Pressure 0 Capillary Temperature 270

3. Set the Multiplier Gain to 300,000. 4. Set the instrument to positive ion mode. 5. Infuse Ultramark 1621 at a flow rate of 1 to 3 µL/min. In Full Scan mode the spectrum should be similar to the

following figure.

6. In the Define Scan view set the parameter values so that they are the same as in the following figure.

7. Choose the Q1 page of the Device view. 8. Select Tube Lens Offset, click Optimize, then click Accept. 9. Select Lens 1-2 Offset, then click Optimize, then click Accept. 10. Select Lens 2-1 Offset, then click Optimize, then click Accept.

FD Ayres 16 of 21 70111-97117 rev C

11. Choose the Q3 page of the Device view. 12. Select Tube Lens Offset, then, click Optimize, then click Accept. 13. Select Lens 3-3 Offset, then click Optimize, then click Accept. 14. Repeat steps 6 to 13 for Ultramark mass 1821.950. The signal intensity must be >1.5 × 105 counts.

15. Choose the Q1 page of the Device view. 16. Select Q1 DC Offset. Add -0.2 to the value for mass 997.389 and enter the sum (-2.8 + -0.2 = -3.0) in the value

for mass 1821.950 as shown in the following figure, then click Apply.

26. Choose the Q3 page of the Device view. 27. Select Q3 DC Offset. Add -0.2 to the value for mass 997.389 and enter the sum in the value for mass

1821.950, then click Apply. 28. In the System Tune and Calibration Workspace choose Ultramark Pos. Ions in the Compound list box. 29. Click Mass Calib for both Q1 and Q3, then click Start. 30. Do not copy the tune table to negative ion mode. 31. Click Save Calib to automatically save the calibration file as the Current Calibration. You will then be prompted

to save the tune file. Save the tune file as Ultramark Pos Ions.TSQTune. 32. Open the Diagnostics Workspace. Then, choose File | Save Calibration As Default Factory. 33. Switch to negative ion mode. 34. Infuse Ultramark at a flow rate of 1 to 3 µL/min. In Full Scan mode the spectrum should be similar to the

following figure.

FD Ayres 17 of 21 70111-97117 rev C

35. In the Define Scan view set the parameter values so that they are the same as in the following figure.

36. Choose the Q1 page of the Device view. 37. Select Tube Lens Offset, then, click Optimize, then click Accept. 38. Select Lens 1-2 Offset, then click Optimize, then click Accept. 39. Select Lens 2-1 Offset, then click Optimize, then click Accept. 40. Choose the Q3 page of the Device view. 41. Select Tube Lens Offset, then, click Optimize, then click Accept. 42. Select Lens 3-3 Offset, then click Optimize, then click Accept. 43. Repeat steps 25 to 32 for Ultramark mass 1879.960. The signal intensity must be >1.5 × 105 counts.

44. Choose the Q1 page of the Device view. 45. Select Q1 DC Offset. Add +0.2 to the value for mass 995.383 and enter the sum in the value for mass

1879.960, then click Apply.

46. Choose the Q3 page of the Device view. 47. Select Q3 DC Offset. Add +0.2 to the value for mass 995.383 and enter the sum in the value for mass

1878.960, then click Apply. 48. In the System Tune and Calibration view choose Ultramark Neg. Ions in the Compound list box. Enter the

masses 1479.980 and 1879.960 (these values will replace the masses 1405.950 and 1805.920) as show in the following figure.

49. Click Mass Calib for both Q1 and Q3, then click Start. 50. Do not copy the tune table to positive ion mode. 51. Click Save Calib to automatically save the calibration file as the Current Calibration. You will then be prompted

to save the tune file. Save the tune file as Ultramark Neg Ions.TSQTune. 52. Open the Diagnostics Workspace. Then, choose File | Save Calibration As Default Factory.

FD Ayres 18 of 21 70111-97117 rev C

Masses for Ultramark + ions Masses for Ultramark - ions 822.0162 880.0228 922.0098 980.0164 1022.0034 1080.0100 1121.9970 1180.0036 1221.9906 1279.9972 1321.9843 1379.9908 1421.9779 1479.9844 1521.9715 1579.9781 1621.9651 1679.9717 1721.9587 1779.9653 1821.9523 1879.9589 1921.9459 1979.9525 2021.9395 2079.9461 2121.9332 2179.9397

FD Ayres 19 of 21 70111-97117 rev C

Installation Procedure for AM Instruments Accurate Mass Performance Specification A mixture of polyethylene glycols (PEGs) of average molecular weights 200, 400, 600, and 1000 is introduced. There are 27 ammoniated PEG ions of interest in the resulting spectrum from 124 Da to 1268 Da. Accurate mass data is generated on each of the 25 ions from 168 Da to 1224 Da using the neighboring peaks as internal lock masses. The mass of each ion is determined from the average of up to 100 scans and the error between the expected mass and the measured mass is expressed in mmu and ppm. The root mean square (RMS) average is computed from the errors of the 25 individual ions. The RMS error will be less than or equal to 5 ppm. The TSQ Vantage AM is calibrated for higher masses by performing a mass calibration followed by an AM calibration and then saving the calibration file. The procedure uses PEG mixture as the calibrant.

Mass Calibrate 1. Set the instrument to Profile mode. 2. Set the Ion Source parameters to values specific to run Mass Calib:

Spray Voltage 4000 Sheath Gas Pressure 2 Aux Gas Pressure 0 Capillary Temperature 270

3. Set the Multiplier Gain to 300,000. 4. Infuse 1,3,6-polytyrosine at a flow rate of 1 to 2 µL /min. 5. Confirm that the spray is stable. Run System Evaluation – API Stability Test. The results must be excellent. 6. Open the System Tune & Calibration Workspace. Select Polytyrosine-1,3,6 from the Compound list box. Run

Mass Calib for both Q1 and Q3. 7. Click Save Calib to automatically save the calibration file as the Current Calibration. You will then be prompted

to save the tune file. Save the tune file as Polytyrosine Accurate Mass.TSQTune. 8. Open the Diagnostics Workspace. Then, choose File | Save Calibration As Default Factory.

Calibrate for High Resolution 1. Run HighRes for both Q1 and Q3. 2. Click Save Calib to automatically save the calibration file as the Current Calibration. You will then be prompted

to save the tune file. Save the tune file as Polytyrosine High Resolution.TSQTune. 3. Open the Diagnostics Workspace. Then, choose File | Save Calibration As Default Factory.

Calibrate for Accurate Mass 1. Infuse the 100 pmol/µL PEG Calibration Solution at a flow rate of 20 µL/min. Flush the system for at least

5 minutes. Confirm that polytyrosine is absent from the spectrum. 2. Confirm that the spray is stable. Run System Evaluation – API Stability Test. The results must be excellent. 3. Check that the peaks m/z 168.1, 520.3, and 1004.6 are stable in both Q1MS and Q3MS. 4. Open the Diagnostics Workspace. Run Accurate Mass Procedures – Linearize System Masses for Q3 SIM.

Save the calibration file. Evaluate the results. There should be no points that lie outside of the calibration curve.

5. Run Accurate Mass Procedures – Check Accurate Mass for Q3. The test result should be better than ±5 ppm RMS.

6. Run Accurate Mass Procedures – Linearize System Masses for Q1 SIM. Save the calibration file. Evaluate the results. There should be no points that lie outside of the calibration curve.

7. Run Accurate Mass Procedures – Check Accurate Mass for Q1. The test result should be better than ±5 ppm RMS.

FD Ayres 20 of 21 70111-97117 rev C

Recipes 200 fg/µL Reserpine Specification Solution 1. Prepare the 2 mg/mL reserpine solution:

a. Add 100 mg of reserpine to a 50 mL volumetric flask. b. Fill the 50 mL flask to volume with methylene chloride. Mix the contents thoroughly.

2. Prepare a 20 ng/µL reserpine solution: a. Use a volumetric pipette to transfer 1 mL of the 2mg/mL solution to a 100 mL volumetric flask. b. Fill the 100 mL flask to volume with methylene chloride. Mix the contents thoroughly.

3. Prepare a 200 pg/µL reserpine solution: a. Use a volumetric pipette to transfer 1 mL of the 20 ng/µL solution to a 100 mL volumetric flask. b. Fill the 100 mL flask to volume with acetonitrile. Mix the contents thoroughly.

4. Prepare a 2 pg/µL reserpine solution: a. Use a volumetric pipette to transfer 1 mL of the 200 pg/µL to a 100 mL volumetric flask. b. Fill the 100 mL flask to volume with acetonitrile. Mix the contents thoroughly.

5. Prepare the 200 fg/µL reserpine solution: a. Use a volumetric pipette to transfer 10 mL of the 2 pg/µL solution to a 100 mL volumetric flask. b. Fill the 100 mL flask to volume with a solution of (70:30) acetonitrile/water. Mix the contents thoroughly.

Ultramark 1621 Calibration Solution 1. Use a syringe to add 100 µL of Ultramark 1621 to a 100 mL volumetric flask. 2. Fill the 100 mL flask to volume with acetonitrile. Mix the contents thoroughly. 3. Use a volumetric pipette to transfer 5 mL of the solution from step 2 to a 100 mL volumetric flask. 4. Use a syringe to add 500 µL of glacial acetic acid to the 100 mL volumetric flask. 5. Fill the 100 mL flask to volume with a mixture of (50:50) methanol/water. Mix the contents thoroughly.

PEG Calibration Solution 1. Prepare the 1000 pmol/µL PEG stock solution:

a. Weigh 20 mg of PEG 200, 40 mg of PEG 400, 60 mg of PEG 600, and 100 mg of PEG 1000 into a clean 100 mL volumetric flask.

b. Fill the 100mL volumetric flask to volume with a mixture of (50:50) methanol/water. Mix the contents thoroughly. 2. Prepare the 100 pmol/µL PEG calibration solution:

c. Use a volumetric pipette to transfer 10 mL of the 1000 pmol/µL solution into a 100 mL volumetric flask. d. Add 154.0 mg of NH4OAc to the volumetric flask. e. Fill the 100 mL volumetric flask to volume with a mixture of (50:50) methanol/water. Mix the contents

thoroughly.

FD Ayres 21 of 21 70111-97117 rev C

Deliverables for the Customer Installation Service Report Included Printed diagnostic test results for System Control Included Printed diagnostic test results for Ion Source Board Included Printed diagnostic test results for RF System Included Printed diagnostic test results for Analyzer Control Board Included Printed calibration report for positive ion mode Included Printed calibration report for negative ion mode (if it is required) Included Printed S/N report that show the instrument passes the performance specification for ESI mode Included Printed S/N report that show the instrument passes the performance specification for APCI mode Included TSQ Series Installation Procedure P/N 70111-97117 Included

Service Engineer Signature

Date