LC/MS Purification System TRILUTION LC v3.0 User s Guide

Transcription

1 LC/MS Purification System TRILUTION LC v3.0 User s Guide Gilson, Inc. PO Box Middleton, WI , USA Tel: or Fax: Gilson S.A.S. 19, avenue des Entrepreneurs BP 145 F Villiers-le-bel, FRANCE sales@gilson.com service@gilson.com training@gilson.com LT Gilson, Inc. All rights reserved.

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3 Table of Contents 1 Introduction System Overview Standard Equipment Required Accessories Optional Accessories Customer Service Repair and Return Policies Before Calling Us Warranty Repair Warranty for Non-Gilson Equipment Non-Warranty Repair Return Procedure Unit End-of-Life Safety Operation Overview LC/MS Method Setup Configuration Start Method Stop Method Injection/Fraction Collection Method Shutdown Method Prepare for a Run Dilute the Tune Mix Fill the Calibration Vials or Syringe Adjust the Probe Position Auto Tune the Flexar SQ 300 MS Detector Check Tune Start the Run Ensure that Detector Lamps are On Set Up the Splitter Set Up the Sample List During the Run View Results and Spectral Data Open Results View Fraction Spectra Browse Analysis for Full Scan MS Channel View Spectral Data Build a Mass Spectral Library Search for Mass Spectral Matches View Spectral Data for Another Sample Fraction Collection Simulation How to Simulate Fraction Collection by Slope or Level View Diagnostics View Status Manage Maintenance Alarms Create an Alarm Modify an Alarm Rename an Alarm Alarm Reminder Maintenance Periodic Maintenance Periodic ESI Source Cleaning Replace the Probe Needle Maintenance of the Roughing Pump Gas Ballasting the Roughing Pump Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide Table of Contents-1

4 A Splitter Connections and Control B Source Change Electrical Connections A-2 Make Connections A-2 Control A-3 Replace ESI Source with APCI Source B-2 Remove the ESI Source B-2 Install the APCI Source B-5 Replace APCI Source with ESI Source B-8 Remove the APCI Source B-8 Install the ESI Source B-10 C Replacement Parts Table of Contents-2 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

5 Introduction 1 The Gilson LC/MS Purification System enables scientists to use the specificity of MS to improve the efficiency of their purification processes and reduce post-purification bottlenecks. The addition of an MS detector allows for more targeted purification, resulting in fewer fractions to be processed, while simultaneously providing MS data. Installation should be performed by your local Gilson representative. This user s guide describes how to operate the system using TRILUTION LC Software v3.0. For additional information, refer to the documentation supplied with each individual instrument and the software. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 1-1

6 Introduction 1 System Overview System Overview The Gilson LC/MS Purification System will arrive in three shipments: one shipment will contain the LC system and accessories, another shipment will contain the MS detector, roughing pump, and accessories, and another shipment will contain the tune mix. Keep the original containers and packing assemblies if components need to be returned. The Gilson LC/MS Purification System uses a GX-281 Prep, GX-271 Prep, or 215 Liquid Handler on the front end. It uses an injector, HPLC pumps, UV detector, splitter, 307 Make Up Pump, and Flexar SQ 300 MS Detector. It is controlled by TRILUTION LC Software v3.0. Standard Equipment Flexar SQ 300 MS Detector Part Number Description FLEXAR SQ 300 MS DETECTOR WITH SYRINGE AND DIVERTER VALVE - USB Accessories Part Number Description ACCESS,LCMS PE SQ300 USB includes: PLMB PKG,LCMS PE SQ PUMP CTL MODULE, (110/220V) PUMP HEAD, 5SC, 5 ML/MIN 8700 PSI SPLITTER, RHEODYNE MRA POWER CORD DETACHABLE, 220V POWER CORD, 18/3, 110V GEARS MRA SPLITTER CONTROL TRILUTION LCMS LICENSE, LIFETIME FLASHLIGHT, 9LED, GILSON 1-2 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

7 Introduction 1 Documentation Part Number Description LT180002CD DOC CD, LCMS PURIFICATION SYSTEM LT PLUMBING DIAGRAM FOR LCMS PE SQ300 LT IQ PROCEDURE TRILUTION LC V3.0 LCMS LT INSTRUCTIONS GEARS MRA SPLITTER CONTROL System Overview Required Accessories Some required accessories must be ordered separately. TRILUTION LC v3.0 Part Number Description R30 TRILUTION LC MEDIA V TRILUTION LC LICENSE, LIFETIME Tune Mix Part Number Description ESI TUNE MIX, AGILENT, 100 ML Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 1-3

8 Introduction 1 System Overview Optional Accessories Optional accessories can be ordered separately. APCI Source For most classes of compounds ESI is the most versatile source to use; however, when using normal phase solvents and highly non-polar compounds, APCI should be considered. APCI is also a more forceful ionization technique and can be used with compounds that do not ionize well with ESI. Part Number Description SQ 300 APCI SOURCE ASSY USB Nitrogen Generator* Due to the high consumption of nitrogen, the use of a nitrogen gas generator is recommended. Gilson offers models specifically designed to meet the needs of the Flexar SQ 300 MS Detector. Part Number Description GEN,NITROGEN PE 120V 35L/MIN GEN,NITROGEN PE 230V 35L/MIN *When using an APCI source, higher nitrogen purity is required. The nitrogen generator cannot be used. Probe The interchangeable, quick-connect probe enables each user to have his own nozzle, preventing cross contamination between multiple user samples and preventing plugging issues from one user to the next. Part Number Description PROBE,MS PE ESI KIT W/CAPILLARY CVR PEMS Roughing Pump Housing To lower the noise from the roughing pump, Gilson offers an acoustically insulated pump box for purchase. Part Number Description HSG,MS PE ROUGH PUMP ACOUSTIC 1-4 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

9 Introduction 1 Customer Service Gilson, Inc. and its worldwide network of representatives provide customers with the following types of assistance: sales, technical support, applications, and instrument repair. If you need assistance, please contact your local Gilson representative. Specific contact information can be found at To help us serve you quickly and efficiently, please refer to Before Calling Us. Customer Service Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 1-5

10 Introduction 1 Repair and Return Policies Repair and Return Policies Refer to the following information and then contact your local Gilson representative. Specific contact information can be found at Before Calling Us Your local Gilson representative will be able to serve you more efficiently if you have the following information: serial number, model number, and version of the instruments involved list of concise symptoms list of operating procedures and conditions you were using when the problem arose list of all instruments in the configuration and the connections to those instruments list of other electrical connections in the room report of diagnostics (refer to View Diagnostics) Warranty Repair Units covered under warranty will be repaired and returned to you at no charge. If you have any questions about applicability, contact your local Gilson representative. Warranty for Non-Gilson Equipment Contact your local Gilson representative. Gilson, Inc. will coordinate with the manufacturer for handling warranty-related service. The warranties are those provided by the manufacturer. Non-Warranty Repair For out-of-warranty repairs, contact your local Gilson representative who will discuss service options with you and can assist in making arrangements to return the equipment, if necessary. 1-6 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

11 Introduction 1 Return Procedure Contact your local Gilson representative to obtain authorization before returning any Gilson equipment. To return a piece of equipment: Carefully pack the unit to prevent damage in transit. Check with your local Gilson representative regarding proper method of shipment. No responsibility is assumed by Gilson or your local Gilson representative for damage caused by improperly packaged instruments. Indicate the authorization on the carton and on the packing slip. Always insure for the replacement value of the unit. Include a description of symptoms, your name, address, phone number, and purchase order to cover repair costs, return and shipping charges, if your organization requires it. Unit End-of-Life When a unit reaches the end of its useful life, refer to the following websites for directions and information on the end-of-life policy: (for Gilson products) or (for PerkinElmer products) This is in accordance with the European Union Directive 2002/96/EC on Waste Electrical and Electronic Equipment (WEEE). Repair and Return Policies Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 1-7

12 Introduction 1 Safety Safety For safety information, refer to the Flexar SQ 300 MS Detector Safety and Compliance Information document supplied on the LC/MS Purification System documentation CD. 1-8 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

13 TRILUTION LC Software provides programmed control of the Flexar SQ 300 MS Detector during a chromatographic run. For each injection made during the run, up to eight channels of data can be acquired and viewed in TRILUTION LC. After the run, a chromatogram at any mass and a spectrum at any time point can be displayed for saved spectra. Additionally, the iso-electric or 3D plot for collected data can be displayed. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-1

14 Overview Overview This chapter provides an overview of the workflow to control the Gilson LC/MS Purification System (including a Flexar SQ 300 MS Detector) in a chromatographic run using TRILUTION LC Software. It assumes that the LC/MS Purification System has been installed by your local Gilson representative. LC/MS Method Setup describes setting up suggested methods. These include a start up method, an injection/fraction collection method using conditional logic fraction collection, a stop method to shut down the system if an error is encountered, and a shutdown method to place the Flexar SQ 300 MS Detector into standby. Prepare for a Run describes how to prepare for a run, adjust the probe position (if necessary), prepare the tune mix and then fill the calibration vials or syringe in preparation for an auto tune or a check tune, use the Mass Spec Auto Tune task to tune the Flexar SQ 300 MS Detector and Mass Spec Check Tune task to verify the last auto tune that was run, view the results of an auto tune or a check tune, and what to do if there is a failure. Start the Run describes how to set up a sample list and start the run. During the Run describes how to view the background subtracted mass spec fraction spectra as it is collected. View Results and Spectral Data describes how to view spectral data obtained from the run. View Diagnostics describes how to check the status of the Flexar SQ 300 MS Detector, if desired. 2-2 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

15 LC/MS Method Setup Example results are provided for the Gilson LC/MS Purification System with TRILUTION LC Software v3.0. There is an example for an ESI application and an example for an APCI application. The methods used to generate those results can be extracted. Gilson recommends using similar methods. The general method setup rules still apply when creating LC/MS methods, which include: Separate tasks in a method 0.02 min before the Start Data Collection task and 0.1 min after the Start Data Collection task. Separate tasks to the same instrument by a minimum of 0.1 min. Use up to, but no more than, eight channels for data collection. Configuration The LC/MS Purification System requires the Flexar SQ 300 MS Detector and the 307 Make Up Pump. The splitter is controlled by contact signals triggered by TRILUTION LC or by setting it up as a GEARS device. It is recommended to designate a UV channel as the primary channel (the channel the software monitors when collecting fractions); however, an MS channel can be designated as the primary channel. LC/MS Method Setup Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-3

16 LC/MS Method Setup Flexar SQ 300 MS Detector The Flexar SQ 300 MS Detector will be detected in a scan. When adding the Flexar SQ 300 MS Detector to the configuration: 1 Select Use next to each MS data channel that will be used. The maximum number of channels per configuration that can be used is eight (seven MS and one UV or six MS and two UV, etc.). 2 Optionally, type a unique description for each channel. If known, the channel description (SIM, Target, Scan, etc.) is recommended. 3 Select the Source Type: Electrospray or APCI. 4 When selecting the data rate, consider the scan range that will be used and refer to the following table: Recommended Maximum Scan Range (High m/z - Low m/z) Data Rate (Data points per second) Scan without SIM Channels Scan with SIM Channels m/z 242 m/z 5 (Default) 714 m/z 515 m/z m/z 1321 m/z 5 On the Units tab, optionally set up user units. Data is scaled, plotted, and analyzed according to these units. All calculations (i.e., heights and areas) are reported according to these units. Units are shown on the graph axis while running and in reports. 6 On the General tab, optionally enter a different instrument name that will be used to identify the Flexar SQ 300 MS Detector in a task. (The default instrument name is MS Detector.) 7 The Flexar SQ 300 MS Detector has a fixed serial number; therefore, the field in the software does not allow editing. 2-4 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

17 307 Make Up Pump When scanning for a 307 Make Up Pump, you will be prompted to select the instrument type. Select 307 Make Up Pump, and then click OK. When adding the 307 Make Up Pump to the configuration: 1 Select the head size. The default is 5, which is correct for what is being supplied with the Gilson LC/MS Purification System. 2 Select the solvent and set the refill speed. LC/MS Method Setup 3 On the Contacts tab, optionally name any contacts that will be used. 4 On the General tab, optionally enter a different instrument name that will be used to identify the 307 Make Up Pump in a task. (The default instrument name is Make Up Pump.) 5 Confirm that the GSIOC ID setting matches the setting on the make up pump and is unique in the system. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-5

18 LC/MS Method Setup Start Method Write a start method to get the LC/MS Purification System ready for a run. Schedule timed tasks to ramp the mobile phase gradient, start the Flexar SQ 300 MS Detector and wait for it to reach proper operating temperature, home the liquid handler, prime, start the make up pump, and ready the splitter for operation. 2-6 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

19 Flexar SQ 300 MS Detector Schedule the Mass Spec Single Quad Start Up task near the beginning of the method. Select the Source Type: ESI or APCI. The default values are acceptable for most applications. Schedule this task a minimum of five minutes BEFORE the Start Make Up Pump task. This allows the Flexar SQ 300 MS Detector to reach proper operating temperature before running samples. LC/MS Method Setup APCI Source When using an APCI source, schedule the Mass Spec Single Quad APCI Settings task in the Start method. This task changes the corona currents used when collecting data. The default settings are acceptable for most applications. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-7

20 LC/MS Method Setup 307 Make Up Pump Schedule the Start Make Up Pump task near the end of the method. The recommended make up pump flow rate for most applications is 0.5 ml/min or less (maximum of 1.4 ml/min). If the make up pump flow rate is greater than or equal to 0.5 ml/min, move the probe and spray away from the source using small increments. Refer to Adjust the Probe Position for instructions. Schedule this task a minimum of five minutes AFTER the Mass Spec Single Quad Settings task. This allows the Flexar SQ 300 MS Detector to reach proper operating temperature before running samples. Splitter Schedule the Contact Open-Close task to close a specified output contact on the liquid handler, which readies the splitter for operation. Use output 2 on the GX-271 Prep or 215 Liquid Handler Use output 3 on the GX-281 Prep Liquid Handler Note: To control the splitter as a GEARS device, instead of by contact control, use the GEARS MRA Splitter Control (part number ). Refer to the documentation supplied with it for more information. 2-8 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

21 Stop Method Write a stop method to shut down the system if an error is encountered. Schedule timed tasks to stop flow from the mobile phase pumps, home the liquid handler, stop the make up pump and the splitter, and put the Flexar SQ 300 MS Detector into standby. When saving this method, be sure to select Use for Error Handling. LC/MS Method Setup Splitter Schedule the Contact Open-Close task near the beginning of the method to close a specified output contact on the liquid handler, which stops the splitter. Use output 2 on the GX-271 Prep or 215 Liquid Handler Use output 3 on the GX-281 Prep Liquid Handler Note: To control the splitter as a GEARS device, instead of by contact control, use the GEARS MRA Splitter Control (part number ). Refer to the documentation supplied with it for more information. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-9

22 LC/MS Method Setup 307 Make Up Pump Schedule the Stop Make Up Pump task near the beginning of the method to stop flow from the 307 Make Up Pump. Flexar SQ 300 MS Detector Schedule the Mass Spec Single Quad Standby task at the end of the method (and at approximately five minutes AFTER the Stop Make Up Pump task) to put the Flexar SQ 300 MS Detector into standby. Note: Do not unplug the Flexar SQ 300 MS Detector or switch the Power On/Off switch to Off. Note: Make up pump flow must be at 0 ml/min and the splitter must be off before putting the Flexar SQ 300 MS Detector into standby. Failure to do so may damage the instrument Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

23 Injection/Fraction Collection Method Start with a typical prep injection/fraction collection method that includes a mobile phase gradient, detector settings, prep injection, start/stop data collection, and start/stop fraction collection, and then add tasks to control the Flexar SQ 300 MS Detector, 307 Make Up Pump, and splitter. LC/MS Method Setup Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-11

24 LC/MS Method Setup Control Flexar SQ 300 MS Detector Mass Spec Single Quad Settings Task Schedule the Mass Spec Single Quad Settings task near the beginning of the method. 1 Under Instrument, select MS Detector or the name entered in the configuration. 2 Under Scan Settings, enter the beginning and ending masses in the full scan range. (Range limits m/z.) Refer to the following table when setting the scan range for the Flexar SQ 300 MS Detector. Gilson recommends that the difference between the high end of the scan range and the low end of the scan range not exceed the values below. Recommended Maximum Scan Range (High m/z - Low m/z) Data Rate (Data points per second) Scan without SIM Channels Scan with SIM Channels m/z 242 m/z 5 (Default) 714 m/z 515 m/z m/z 1321 m/z By default, Save Spectra is selected, which means the full scan data will be saved and can be viewed as spectral data. 3 Under Channel, select the MS channel number. 4 Select the type of monitoring for the selected channel number: Scan - All Masses - Select to display the data trace as the total ion count for the masses within the specified scan range (full scan). If Save Spectra was selected (it is by default), the data for the full scan range will be saved and can be viewed as spectral data. Do not associate an analysis with the data channel used for the scan of all masses. An analysis can be browsed in for the channel post-run and re-analyzed. Scan - Target Mass - Target - Select to display the data trace as the specified mass with one to three adducts (extracted ion or extracted ions). The specified mass with adducts must be within the scan range. If Save Spectra was selected (it is by default), the data for the full scan range will be saved and can be viewed as spectral data. SIM - Target - Select to monitor any target mass with one to three adducts (Selected Ion Monitoring). The specified mass with adducts must be within the range limits ( m/z) Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

25 5 If Scan - Target Mass - Target or SIM - Target was selected: a) Enter the Target mass value. b) Enter values for one to three adducts. Select the box for adducts to be used and then enter a value for each. Note: The signal intensities for the selected adducts for a specified channel are summed. This summed signal is the one displayed and used for fraction collection. This is done to mitigate fluctuations to the channel signal due to adduct distribution factors. 6 Under Data Alignment, enter a time value to shift the retention time of the MS peak to align with the UV peak. Only positive values are allowed, which shifts the MS data trace to the right. The following example shows how use of the data alignment feature can align your UV and MS peaks for optimal fraction collection. Using the crosshair tool, determine the time at the apex of the peak that elutes later (UV) and then subtract the time at the apex of the peak that elutes first (MS). The difference is the data alignment time. If the UV peak elutes first, try making the following system adjustments: Add tubing between the splitter and the UV flow cell inlet. Shorten tubing between the splitter and the Flexar SQ 300 MS Detector Increase the make up pump flow rate. Note: This can cause MS peaks to become wider and less intense. LC/MS Method Setup A B Data Alignment Time = 0 min Data Alignment Time = 0.1 min Data Alignment Time = Time at Apex of Peak B - Time at Apex of Peak A Note: If the primary channel (for fraction collection) is an MS channel, this value must be set to 0 (which is the default). Additionally, the value for Fraction Collection Delay Volume will need to be calculated and then entered on the liquid handler configuration property page using the following equation: Fraction Collection Delay Volume = Fraction Collection Delay Volume (determined by your local Gilson representative) + [Time needed to align MS peaks with UV peaks * Mobile Phase Flow Rate] Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-13

26 LC/MS Method Setup 7 Under Polarity, select to form positive ions or negative ions on all channels. When forming positive ions, adduct values are added to the target. When forming negative ions, adduct values are subtracted from the target. 8 Repeat steps 3 5 for any additional channels (up to seven). After specifying monitor settings for channels to be used, it is recommended to rename the data channels with the monitor description (SIM, Target, Scan, etc.). Do not associate an analysis with the data channel used for the scan of all masses. An analysis can be browsed in post-run and data re-analyzed. Sync Schedule a Sync task in the method BEFORE the injection task that syncs to the end of the Mass Spec Single Quad Settings task to prevent starting data collection before the MS settings are set. 307 Make Up Pump Start Make Up Pump Although the make up pump flow was started in the start method, the Start Make Up Pump task needs to be scheduled in the injection/fraction collection method too. Running the task in this method sets its compressibility. The recommended make up pump flow rate for most applications is 0.5 ml/min or less (maximum of 1.4 ml/min). If the make up pump flow rate is greater than or equal to 0.5 ml/min, move the probe and spray away from the source using small increments. Refer to Adjust the Probe Position for instructions Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

27 Splitter 1 Schedule a Contact Open-Close task BEFORE the injection task and AFTER the Start Make Up Pump task to open a specified output contact on the liquid handler, which starts the splitter. Use output 2 on the GX-271 Prep or 215 Liquid Handler Use output 3 on the GX-281 Prep Liquid Handler LC/MS Method Setup 2 Schedule a Contact Open-Close task at the end of the method to close the contact and stop the splitter. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-15

28 LC/MS Method Setup Conditional Fraction Collection Task The Conditional Fraction Collection task enables setting up conditional logic fraction collection using data on up to eight channels. This is ideal for using both UV and MS detection to trigger collection. This task can be used in place of the Fraction Collection Settings task. 1 Select the primary condition (either slope or level) on the primary channel. Use of a UV channel as the primary channel is recommended. 2 Select the AND option button to indicate that fraction collection will occur when the condition on the primary channel is met AND all secondary conditions are met. Alternatively, select the OR option button to indicate that fraction collection will occur when the condition on the primary channel is met OR any secondary condition is met. Alternatively, select the AND/OR option button to indicate that fraction collection will occur when the condition on the primary channel is met AND any secondary condition is met. 3 Set conditions on up to seven additional channels using the tabs. The text color of the tab number changes from red to green when a channel has been selected and conditions have been set. Each condition could use a different channel or multiple conditions could be set up on the same secondary channel. 4 Optionally, select the Co-eluted Peaks check box to collect co-eluted peaks. 5 Optionally, select the Collect Non Peaks check box to collect non peaks. 6 Select the Fraction Sites tab. a) If using a liquid handler as a fraction collector, optionally set the collection and travel depths. The fraction collection depth is the probe height used when dispensing into fraction sites. The fraction travel depth is the probe height used when moving between fraction sites. The probe will move to the Z-value entered; if 0 is entered, the probe will move to the bed bottom (no safety factor incorporated). The valid range is the Z travel distance and -1, where -1 is equal to the Z Safe Height. b) Select a fraction collector and zone where fraction collection will begin for an injected sample (Fraction Site 1). The default fraction well is 0, the indication for continuous collection where fractions are always collected in the next available tube. c) Select up to three additional fraction collectors and/or zones (Fraction Site 2, Fraction Site 3, and Fraction Site 4). 7 If subdividing, drag the Set Peak Per Tube or Set Non Peak Per Tube task and then drop it in the workspace after the Conditional Fraction Collection task. For non peak collection, select the Collect Non Peaks check box. Subdivision can be by time or volume. 8 Add the Start Fraction Collection (after the Set Peak Per Tube or Set Non Peak Per Tube task) and the Stop Fraction Collection tasks to the method Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

29 Control - Error Handling Flexar SQ 300 MS Detector Optionally, use error handling to check the status of the Flexar SQ 300 MS Detector. Run the stop method if an error is encountered. The Flexar SQ 300 MS Detector status is not updated during data collection. Control error handling parameters for the Flexar SQ 300 MS Detector are described below. APCI Vaporizer Temperature This error results when the APCI vaporizer temperature exceeds or is equal to or is lower than the value set on this screen, depending on the mathematical operator used. Dry Gas Flow This error results when the dry gas flow exceeds or is equal to or is lower than the value set on this screen, depending on the mathematical operator used. Dry Gas Temperature This error results when the dry gas temperature exceeds or is equal to or is lower than the value set on this screen, depending on the mathematical operator used. Instrument Error This error results when any scheduled command fails to execute or if the software loses communication with an instrument. Nebulizer Gas Pressure This error results when the nebulizer gas pressure exceeds or is equal to or is lower than the value set on this screen, depending on the mathematical operator used. Not Pumped Down This error results when the MS Detector is not pumped down. The valid mathematical operator is == and the valid value is TRUE. Standby This error results when the MS Detector goes into standby. The valid mathematical operator is == and the valid value is TRUE. Vented This error results when the MS Detector vents. The valid mathematical operator is == and the valid value is TRUE. Check Tune Area Failure/Check Tune Mass Failure/Wait Time Not used. LC/MS Method Setup Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-17

30 LC/MS Method Setup 307 Make Up Pump The error handling parameters available for the 307 Make Up Pump are described below. Contact Error This error results when an error input contact is activated. The valid mathematical operator is == and the valid values are Open and Closed. Instrument Error This error results when any scheduled command fails to execute or if the software loses communication with an instrument. Pressure This error results when the system pressure exceeds or is equal to or is lower than the value set on this screen. Set up error handling to run a stop method to stop all pumps if the pressure exceeds 1000 psi, as this is the maximum pressure rating for the splitter in the system. Analysis Report Features for reporting MS data are described below. Use of the auto export feature on any channel is NOT recommended when using a Flexar SQ 300 MS Detector. Graph Settings An option for reporting mass spectral data on the report graph is described below. Peak Mass (PM) Peak Mass (PM) Select to label each peak on the chromatogram with its most abundant m/z value Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

31 Spectrum Settings - Mass Spec Spectrum Tab Choose whether spectral views will be displayed and how they will be displayed in the report. Spectral Views - 3D Plot/Iso-electric Plot Select to display the 3D Plot and/or Iso-electric Plot in the report. Optionally, specify elevation angle and rotation for the 3D plot display. Optionally specify stepped intervals, continuous, or grayscale for the 3D and iso-electric plot display. Spectral Views - Minimum Mass (m/z)/maximum Mass (m/z) Enter the range to display both in the report and when viewing spectral data from the results. LC/MS Method Setup Spectrum Size Enter the percentage of the page to use for a spectrum display in the report. Peak Spectral Views - Peak Spectra Select to display the peak spectra for all integrated peaks. Additionally, choose to display the peak spectra at the retention time or display the entire peak spectra. Neither fraction spectra nor peak spectra will be displayed with the Report Template 2 option (Report Template 1 is the default). Peak Spectral Views - Peak Fraction Spectra Select to display the fraction spectra for all integrated peaks. Additionally, choose the number of masses to show. Peak fraction spectra will only be displayed for an analysis associated with the primary channel. Neither fraction spectra nor peak spectra will be displayed with the Report Template 2 option (Report Template 1 is the default). Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-19

32 LC/MS Method Setup Shutdown Method Write a shutdown method to shut down the system. When finished running samples, it is recommended to run a shutdown method. Schedule timed tasks to stop flow from the mobile phase pumps, stop the splitter and make up pump, home the liquid handler, and put the Flexar SQ 300 MS Detector into standby. Splitter Schedule the Contact Open-Close task to close a specified output contact on the liquid handler, which stops the splitter. Use output 2 on the GX-271 Prep or 215 Liquid Handler Use output 3 on the GX-281 Prep Liquid Handler Note: To control the splitter as a GEARS device, instead of by contact control, use the GEARS MRA Splitter Control (part number ). Refer to the documentation supplied with it for more information Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

33 307 Make Up Pump Schedule the Stop Make Up Pump task near the beginning of the method to stop flow from the 307 Make Up Pump. LC/MS Method Setup Flexar SQ 300 MS Detector Schedule the Mass Spec Single Quad Standby task at the end of the method (and at approximately five minutes AFTER the Stop Make Up Pump task) to put the Flexar SQ 300 MS Detector into standby. Note: Do not unplug the Flexar SQ 300 MS Detector or switch the Power On/Off switch to Off. Note: Make up pump flow must be at 0 ml/min and the splitter must be off before putting the Flexar SQ 300 MS Detector into standby. Failure to do so may damage the instrument. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-21

34 Prepare for a Run Prepare for a Run Make up pump flow must be at 0 ml/min and the splitter must be off before putting the Flexar SQ 300 MS Detector into standby. Failure to do so may damage the instrument. Regular tunings are recommended, especially if you notice changes in performance. At minimum, it is recommended to run a check tune daily. A check tune is run automatically after the auto tune. Store check tune results in a folder for tracking of long-term trends. A calibrant can be introduced into the Flexar SQ 300 MS Detector by several methods including: using the calibration vials that are located on the front, using the on-board syringe pump, which is also located on the front (right side), or by using an external syringe pump. This section describes how to prepare for the run and includes instructions for running an auto tune/check tune, or just a check tune. Dilute the Tune Mix The recommended tune mix is Electrospray Calibrant Solution, 100 ml (part number ). Gilson recommends diluting this tune mix 1:10 in 95%/5% (v/v) Acetonitrile/Water, using HPLC grade solvents (LCMS grade is recommended). Fill the Calibration Vials or Syringe Calibration Vials There are two calibration vials: vial 1 is on the left and vial 2 is on the right. Either can be used for auto tuning, as there is a selection for each in the Mass Spec Auto Tune task. 1 To remove a calibration vial, reach in and unscrew it. 2 Fill the vial with approximately 15 ml of diluted tune mix. 3 Re-install the calibration vial, ensuring that the small draw tube is inserted into the calibration vial and that the calibration vial is securely fastened into the blue cap to avoid pressure leaks. Note: BOTH calibration vials must be installed, even if only one will be used, as both are pressurized for calibration delivery from a common nitrogen source inside the Flexar SQ 300 MS Detector Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

35 Syringe The on-board syringe pump can be used to deliver the diluted tune mix to the probe for an auto tune or check tune. Remove the Syringe If the syringe is installed, remove it to fill it with diluted tune mix. 1 Pull and twist the spring-loaded thumb knob until the black plate drops. 2 While holding the syringe, pull and turn the black spring-loaded syringe holder. This will free the syringe for removal. spring-loaded thumb knob Prepare for a Run Fill the Syringe 1 Disconnect the tubing to the syringe from the union. 2 Fill the syringe with diluted tune mix. Install the Syringe 1 Reconnect the syringe tubing to the union. 2 While holding the syringe, align the wings on the syringe with the grooves in the syringe holder. 3 While still holding the syringe, turn the black spring-loaded syringe holder and then release it gently. 4 Let go of the syringe and make final positioning adjustments. 5 Move the black plate up to the syringe and then turn the silver thumb knob to lock. black spring-loaded syringe holder Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-23

36 Prepare for a Run Adjust the Probe Position It may be necessary to adjust the probe and spray for any of the following reasons: High sample concentration Move the probe and spray away from the source using small increments Make up pump flow rate is greater than or equal to 0.5 ml/min Move the probe and spray away from the source using small increments Failed auto tune or check tune Ensure that the ESI source is installed, and then refer to the appropriate instructions for the tune mix source to be used. Spray at 300 μl/min For information about how to change the source, refer to Appendix B, Source Change Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

37 Adjust the Probe Position for Auto Tune/Check Tune Use the supplied flashlight to view the spray when making probe adjustments. A good spray is a fine mist that is steady (not pulsing) and is symmetrical. 1 On the Desktop, double-click the TRILUTION LC icon ( ) to start TRILUTION LC. Note: It is imperative that TRILUTION LC is started in this manner when preparing and running an auto tune. 2 In the Project Library, select an application containing a method using a configuration that includes a Flexar SQ 300 MS Detector, and then click Run. The Application Run window appears. 3 Under Method Configuration, select a method using a configuration that includes a Flexar SQ 300 MS Detector (if not already selected), and then click. Prepare for a Run 4 Click to access the Manual Control window. 5 Run the Mass Spec Single Quad Start Up task (in Manual Control). Select ESI. Change the Dry Gas Flow value to 8000 ml/min. Change the Dry Gas Temperature to 300 C. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-25

38 Prepare for a Run 6 Run the Mass Spec Single Quad Settings task (in Manual Control). Set up the scan settings to scan around one of the desired masses. 7 Select the tune mix source. If calibration vial, run the Mass Spec Vials command (in Manual Control). Select the calibration vial to be used: vial 1 is on the left and vial 2 is on the right. If syringe, run the Mass Spec Syringe command (in Manual Control) Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

39 8 Click Chart to enable viewing of the strip chart. 9 Click Start Mass Spec Scan to begin viewing data from Flexar SQ 300 MS Detector Channel 1. The trace for the channel appears in the window. Prepare for a Run 10 Click List and then run the Mass Spec Diverter Valve command. Change Diverter Valve Position to Click Chart to monitor the data trace. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-27

40 Prepare for a Run 12 The marks on the probe assembly can be used for quick positioning. Observe the needle through the inspection window and adjust the tip if necessary. 13 Loosen the lock ring. 14 Turn the needle assembly adjustment screw until the needle tip protrudes about 0.5 mm from the nozzle. 15 Tighten the lock ring. adjustment screw lock ring 16 Adjust the probe position using the black thumb knob on the right. Spinning it up moves the probe to the right. Spinning it down, moves the probe to the left. Monitor the data collection and make adjustments that maximize the peak intensity and signal. However, do not position the probe to spray directly into the capillary entrance, as this can damage the Flexar SQ 300 MS Detector. turn this 2-28 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

41 17 Adjust the probe tilt using the black thumb knob on the left. Spinning it left tilts the probe away from the capillary. Spinning it right tilts the probe towards the capillary. Monitor the data collection and make adjustments that maximize the peak intensity and signal. However, do not position the probe to spray directly into the capillary entrance, as this can damage the Flexar SQ 300 MS Detector. 18 Click Stop Mass Spec Scan. When data collection is halted, a dialog box appears that enables you to save the collected data. turn this Prepare for a Run If you select to save the strip chart, you are prompted to name it, and then it is stored with other run results. The software indicates in the Info Window that the result is a manual result (Manual Result: Yes). 19 Stop the tune mix flow. If a calibration vial was used, run the Mass Spec Vials command (in Manual Control). Select the calibration vial that was used: vial 1 is on the left and vial 2 is on the right. Change the State to OFF. If a syringe was used, run the Mass Spec Syringe command (in Manual Control). Change the Syringe Flow Rate (ml/min) to 0. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-29

42 Prepare for a Run Auto Tune the Flexar SQ 300 MS Detector Auto tune the Flexar SQ 300 MS Detector if changes in performance are observed or if the check tune failed. A check tune is run automatically after the auto tune. 1 Ensure that the ESI source is installed. For information about how to change the source, refer to Appendix B, Source Change. 2 On the Desktop, double-click the TRILUTION LC icon ( ) to start TRILUTION LC. Note: It is imperative that TRILUTION LC is started in this manner when preparing and running an auto tune/check tune. 3 In the Project Library, select an application containing a method using a configuration that includes a Flexar SQ 300 MS Detector and then click Run. The Application Run window appears. 4 Under Method Configuration, select a method using a configuration that includes a Flexar SQ 300 MS Detector (if not already selected), and then click. 5 Click to access the Manual Control window. 6 Run the Mass Spec Single Quad Start Up task (in Manual Control). Change the Dry Gas Flow value to 8000 ml/min. Change the Dry Gas Temperature to 300 C. 7 Wait 5 minutes for the Flexar SQ 300 MS Detector to equilibrate. 8 If samples will be run for positive ionization, continue to step 9. If samples will be run for negative ionization only, skip to step Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

43 9 Run the Mass Spec Auto Tune task (in Manual Control) for positive settings. a) Select the appropriate Tune Mix Source. b) Optionally, select to Prime. (For example, if syringe was refilled after probe positioning.) c) Select Create New Positive Settings. d) Select masses to calibrate. Sample masses should be encapsulated within the selected calibrant masses. All calibrant masses within this range should be selected. Default settings are for use with the tune mix Electrospray Calibrant Solution, 100 ml (part number ), which must be diluted 1:10 in 95%/5% (v/v) Acetonitrile/Water. A check tune will run automatically following the auto tune. 10 View the Check Tune Results. (The dialog appears automatically when the check tune completes.) The Auto Tune Log that appears by default is the log that was generated during the auto tune/check tune. Select Check Tune Results to display the results of the check tune that was run automatically after the auto tune. If any failures for Mass or Width are indicated, first verify the following and then auto tune again: Was the correct tune mix source selected and is there enough tune mix available? Are the lines primed? Are there any leaks in the fittings for the tune mix or nitrogen supply connections? Is the dry gas flow at 8000 ml/min? Is the dry gas temperature at 300 C? Is the nebulizer gas pressure at 80 psi? (Run the diagnostics or check the status box in the Application Run window.) Is there a good spray? (Use the supplied flashlight to view the spray. A good spray is a fine mist that is steady and symmetrical.) 11 If samples will be run for positive ionization only, skip the remaining steps in this procedure. Prepare for a Run Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-31

44 Prepare for a Run 12 Run the Mass Spec Auto Tune task (in Manual Control) for negative settings. a) Select the appropriate Tune Mix Source. b) Optionally, select to Prime. (For example, if syringe was refilled after running the first auto tune.) c) Select Create New Negative Settings. d) Select masses to calibrate. Sample masses should be encapsulated within the selected calibrant masses. All calibrant masses within this range should be selected. Default settings are for use with the tune mix Electrospray Calibrant Solution, 100 ml (part number ), which must be diluted 1:10 in 95%/5% (v/v) Acetonitrile/Water. A check tune will run automatically following the auto tune. 13 View the Check Tune Results. (The dialog appears automatically when the check tune completes.) The Auto Tune Log that appears by default is the log that was generated during the auto tune/check tune. Select Check Tune Results to display the results of the check tune that was run automatically after the auto tune. If any failures for Mass or Width are indicated, first verify the following and then auto tune again: Was the correct tune mix source selected and is there enough tune mix available? Are the lines primed? Are there any leaks in the fittings for the tune mix or nitrogen supply connections? Is the dry gas flow at 8000 ml/min? Is the dry gas temperature at 300 C? Is the nebulizer gas pressure at 80 psi? (Run the diagnostics or check the status box in the Application Run window.) Is there a good spray? (Use the supplied flashlight to view the spray. A good spray is a fine mist that is steady and symmetrical.) 2-32 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

45 Check Tune 1 Ensure that the ESI source is installed. For information about how to change the source, refer to Appendix B, Source Change. 2 Start TRILUTION LC. 3 Under Method Configuration, select a method using a configuration that includes a Flexar SQ 300 MS Detector (if not already selected), and then click. 4 Click to access the Manual Control window. 5 Run the Mass Spec Single Quad Start Up task (in Manual Control). Change the Dry Gas Flow value to 8000 ml/min. Change the Dry Gas Temperature to 300 C. 6 Wait 5 minutes for the Flexar SQ 300 MS Detector to equilibrate. 7 If samples will be run for positive ionization, continue to step 8. If samples will be run for negative ionization only, skip to step 11. Prepare for a Run Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-33

46 Prepare for a Run 8 Run the Mass Spec Check Tune task (in Manual Control) for positive settings. Select the appropriate Tune Mix Source. Optionally, select to Prime. (For example, if syringe was refilled after probe positioning.) Select Positive for the Tune Mix Polarity. 9 View the Check Tune Results. (The dialog appears automatically when the check tune completes.) The Check Tune Log that appears by default is the log that was generated during the check tune. Select Check Tune Results to display the results of the check tune. If any failures for Mass or Width are indicated, first verify the following and then check tune again. If the check tune fails again, run an auto tune. Was the correct tune mix source selected and is there enough tune mix available? Are the lines primed? Are there any leaks in the fittings for the tune mix or nitrogen supply connections? Is the dry gas flow at 8000 ml/min? Is the dry gas temperature at 300 C? Is the nebulizer gas pressure at 80 psi? (Run the diagnostics or check the status box in the Application Run window.) Is there a good spray? (Use the supplied flashlight to view. 10 If samples will be run for positive ionization only, skip the remaining steps in this procedure Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

47 11 Run the Mass Spec Check Tune task (in Manual Control) for negative settings. Select the appropriate Tune Mix Source. Optionally, select to Prime. (For example, if syringe was refilled after running the first auto tune.) Select Negative for the Tune Mix Polarity. 12 View the Check Tune Results. (The dialog appears automatically when the check tune completes.) The Check Tune Log that appears by default is the log that was generated during the check tune. Select Check Tune Results to display the results of the check tune. If any failures for Mass or Width are indicated, first verify the following and then check tune again. If the check tune fails again, run an auto tune. Was the correct tune mix source selected and is there enough tune mix available? Are the lines primed? Are there any leaks in the fittings for the tune mix or nitrogen supply connections? Is the dry gas flow at 8000 ml/min? Is the dry gas temperature at 300 C? Is the nebulizer gas pressure at 80 psi? (Run the diagnostics or check the status box in the Application Run window.) Is there a good spray? (Use the supplied flashlight to view. Prepare for a Run Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-35

48 Start the Run Start the Run Ensure that Detector Lamps are On Lamps on both the 15X Detectors and 171/172 Diode Array Detectors require a minute warm up time. Ensure that the lamps are on before setting up the run. Refer to the detector user s guide for more information. Set Up the Splitter Refer to Appendix A, Splitter Connections and Control for information about how to set up the splitter for a run. Note: To control the splitter as a GEARS device, instead of by contact control, use the GEARS MRA Splitter Control (part number ). Refer to the documentation supplied with it for more information. Set Up the Sample List In the Application Run window, set up the sample list. 1 Insert the start method as the first sample. Setting the Report column option to NO for this sample is recommended, as there is no injection to display in the analysis report. 2 Insert the injection/fraction collection method for as many samples as desired to run. Note: The software will notify you of a configuration change. This is expected as configurations were given unique names. Click OK. 3 Insert the shutdown method as the last sample. Setting the Report column option to NO for this sample is recommended, as there is no injection to display in the analysis report. 4 Satisfy all variables. 5 Optionally, set up any other features available in the Application Run window, if desired. For more information, refer to the TRILUTION LC on-line help or user s guide. 6 Click Run to start running the methods in the application run Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

49 During the Run During the application run, the background subtracted mass spec fraction spectra is displayed as it is collected (if applicable) on the Mass Spec Spectra for Fractions tab. Fraction spectra is obtained from a scan of all masses or a scan with a target mass. The background to be subtracted is determined from averaged spectra at the start of the scan. During the Run Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-37

50 View Results and Spectral Data View Results and Spectral Data Open Results 1 Double-click the results icon ( ) or right-click on an application in the Project Library, and then select Results to open the Results window. 2 Double-click the Run Name to show the Run Results. 3 When the Run Results window appears, the first sample displayed is the Start method. Click Next to display the first sample injection. Traces for the data channels that were marked for use in the configuration are displayed. These are also the traces that were displayed real-time. View Fraction Spectra Double-click on a fraction in the Fraction Bed Layout to display the background subtracted mass spec fraction spectra, if applicable, and to highlight the fraction in the chromatogram Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

51 Browse Analysis for Full Scan MS Channel 1 From the Device/Channel drop-down, select the MS channel that was used for a scan of all masses (as defined in the Mass Spec Single Quad Settings task). 2 Next to the Analysis drop-down, click to open the Browse Analysis window. 3 From the list of analyses, select one. The method(s) in which it is used, the instrument(s) and data channel(s) on which it is used is displayed on the right under Where Used. Choose an analysis from a method that was NOT used to obtain the open run results. 4 Because the selected channel does not currently have an analysis associated with it, the option to Overwrite existing analysis for this sample. is not available. Select Add analysis for all samples using this channel or overwrite., and then click OK. The analysis is now associated with the selected channel for all samples in the open result. 5 On the Run Results window, and with the same channel still selected, click Edit to open the Result Analysis Builder. View Results and Spectral Data 6 On the Peak Integration tab, double-click the icon ( ) for the Analysis Settings integration task to edit the integration parameters. 7 The default Peak Width setting is 0.20 min. When integrating data on a full scan channel, a minimum peak width of 2.0 min is recommended to filter out noise. Edit the peak width value (and slope values and peak filter, if desired) and then click OK. 8 Optionally, name the peaks. 9 On the Report tab, click Graph Settings... Set desired graph options for the report. Select Peak Mass to label each peak in the graph with its most abundant m/z value, and then click OK. 10 Select Spectrum and then Spectrum Settings... Select the Mass Spec Spectrum tab. Set desired spectrum display settings for the report. Select Peak Spectra to display in the report the peak spectra for all integrated peaks. 11 Click OK to close the Result Analysis Builder. 12 Click Re-Analyze to apply the analysis settings. 13 When asked if you want to save data, click Yes. A save completed message will appear. Click OK. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-39

52 View Results and Spectral Data View Spectral Data Using the options in the Spectral Data window, view spectral information obtained from a scan of all masses or a scan with a target mass. 1 From the Device/Channel drop-down, select the MS channel that was used for a scan of all masses or a scan with a target mass (as defined in the Mass Spec Single Quad Settings task). 2 Click Spectral Data. The Spectral Data window appears for viewing chromatogram and spectral plots and creating mass spectral libraries using data collected by the Flexar SQ 300 MS Detector Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

53 Display and Update the Spectrum Plot 1 Display a spectrum. a) In the Spectral Data window, click to select the Spectral Data tab. By default, the spectrum displayed in the upper right is the APEX for: the first named peak in the peak table found or the first integrated peak found (if no peaks were named in the Peak Table) Peak 1 spectrum chromatogram View Results and Spectral Data b) In the Chromatogram toolbar, click to select the cross hair tool ( ), which displays the time and the signal at the intersect of the cross hair. c) Move the mouse pointer to a location within a peak. Notice how the time and signal readings update. Double-click. The spectrum at the apex of the peak appears in the upper-right corner. after double-click, the spectrum plot updates to display the peak 2 Display the spectrum at the APEX for a different peak by double-clicking in the peak on the chromatogram. time and signal readings the cross hair tool is selected and positioned in the peak Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-41

54 View Results and Spectral Data Display and Update the Iso-electric Plot 1 Display the iso-electric plot: a) In the Spectral Data window, click to select the Spectral Data tab. b) Click to show the iso-electric plot. 2 Use the iso-electric plot to update the chromatogram plot and spectrum plot. a) Click to select the cross hair tool ( ), which displays the time, mass, and signal at the intersect of the cross hair. b) Move the mouse pointer over the iso-electric plot. Notice how the time, mass, and signal readings update. Click to update the mass of the overlaid trace on the chromatogram plot and the retention time on the spectrum plot. the cross hair tool is selected time, mass, and signal readings after click, mass of overlaid trace updates after click, spectrum retention time updates c) To better view the trace, optionally hide the trace for the full scan channel: 1) Click to open the Channel Attributes dialog. 2) Select Hide for the full scan channel and then click OK. overlaid trace with the full scan channel hidden 3 Reduce or enlarge the iso-electric plot. To zoom in, right-click on the plot. To zoom out, right-click outside the plot on either axis Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

55 Display and Update the 3D Plot 1 Display the 3D plot. a) In the Spectral Data window, click to select the Spectral Data tab. b) Click to show the 3D plot. Note that the 3D plot is displayed by default. You only need to click the icon if the iso-electric plot is being displayed. 2 Reduce or enlarge the 3D plot. To zoom in, right-click on the plot. To zoom out, right-click outside the plot on either axis. 3D plot View Results and Spectral Data Modify the Displayed Mass Range To modify the mass range displayed in the 3D plot, iso-electric plot, and spectrum plot: 1 If open, close the Spectral Data window by clicking the x in the upper right corner. The Run Results window appears. 2 On the Run Results window, click Edit. The Result Analysis Builder appears. 3 Click to select the Report tab. 4 On the Report tab, select the Spectrum check box and then click Spectrum Settings... The Spectrum Settings dialog appears. Select the Mass Spec Spectrum tab. 5 On the Spectrum Settings dialog, change the Minimum Mass (m/z) and/or the Maximum Mass (m/z), and then click OK. 6 Click OK to exit the Result Analysis Builder. 7 Click Spectral Data to access the Spectral Data window and Spectral Data tab where the 3D plot, iso-electric plot, and spectrum plot are displayed showing the updated mass range. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-43

56 View Results and Spectral Data Build a Mass Spectral Library 1 Create a mass spectral library. a) From the Run Results, click Spectral Data and then select the Library tab. b) Click New to create a new mass spectral library. The New Mass Spectral Library dialog appears. c) In the New Mass Spectral Library dialog: 1) In the Spectral Library Name field, type a name. 2) In the Short Description field (optional), type a brief description of the library. 3) In the Long Description field (optional), type a detailed description of the library. 4) Click OK. The new library opens. 2 Add a peak spectrum into the mass spectral library for the peak. a) Select a peak from the drop-down list of Integrated Peaks. b) Click Add. The Add Mass Spectral Library Entry dialog appears. 3 In the Add Mass Spectral Library Entry dialog: a) In the Peak Name field, change the peak name, if desired. Peak names must be unique to ALL mass spectral libraries. b) In the Comment field (optional), type a brief description of how data for the peak was acquired. For example, include mobile phase and column information. c) In the MS Comparison Parameters (m/z) fields, specify the masses TRILUTION LC will use when comparing the mass spectral library peak against an unknown peak. d) Select the check box next to the Library Name for each mass spectral library to which the peak should be added. e) Click OK to exit the Add Mass Spectral Library Entry dialog and save the peak spectrum. 4 View the spectrum for a peak by clicking the table row in the mass spectral library Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

57 Search for Mass Spectral Matches Post-Run tabbed spectrum view To use run results to search mass spectral libraries: 1 Open Results containing integrated peaks. 2 From the Device/Channel drop-down, select the MS channel that was used for a scan of all masses or a scan with a target mass (as defined in the Mass Spec Single Quad Settings task). 3 From the Run Results, click Spectral Data and then select the Library tab. 4 Select the name or number of one or more integrated peaks from the drop-down list of Integrated Peaks. Only selected peaks can be used when searching for spectral matches. As each peak is selected, a tab appears above the spectrum with the name in red. select integrated peaks here 5 Click Search. The Search Mass Spectral Library dialog appears. 6 In the Search Spectral Library dialog: a) Select the check box next to the Library Name for each library to be searched. b) For each peak spectrum to match, do either of the following: Select the check box next to the Peak Name and under All Peaks to search all mass spectral library peaks. Select the check box next to the Peak Name and under Time Period and then enter Start and End times to limit the search to spectral library peaks whose retention times are within that time period. c) In the Match Threshold field, indicate the smallest match value to be reported. A match value of 1000 indicates a perfect match. 7 Click OK to begin the search. 8 When the search completes, TRILUTION LC displays the five closest matches. 9 In the tabbed spectrum view, select a spectrum. From the drop-down list of Library Matches, select a spectrum to overlay. The integrated peak spectrum appears red, the matched spectrum appears black. 10 Remove the overlaid spectrum by right-clicking on the spectrum tab and then selecting Close Tab to close the spectrum tab. View Results and Spectral Data Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-45

58 View Results and Spectral Data Run Time Optionally, set up the analysis to perform spectral matching on unknown peaks automatically at run time. 1 Open a method. 2 Select the Analysis tab. Mass spectral matching is available for every data channel when mass spectral data is saved, including non-ms channels. When mass spectral matching is performed on a data channel, the saved mass spectral data at the retention time of an integrated peak is compared to mass spectra for peaks stored in the library. 3 Click Mass Spectral Library. The Search Mass Spectral Library dialog appears. 4 In the Search Mass Spectral Library dialog: a) Select the check box next to the Library Name for each library to be searched. b) For each peak spectrum to match, do either of the following: Select the check box next to the Peak Name and under All Peaks to search all mass spectral library peaks. Select the check box next to the Peak Name and under Time Period and then enter Start and End times to limit the search to mass spectral library peaks with retention times within that time period. c) In the Match Threshold field, indicate the smallest match value to be reported. A match value of 1000 indicates a perfect match. d) Click OK to exit the Search Mass Spectral Library dialog. 5 Add mass spectral match columns to the analysis report. a) With the method still open, click to select the Report tab. b) Under Report Type, click to select Sample. c) Under Contents, click to select MS Spectral Match 1. d) Click the right arrow ( ) to add it to the bottom of the Column Contents list. e) Repeat steps c and d to add MS Spectral Match 2 and MS Spectral Match 3 to the list. Note: The Sample Table in the Analysis Report can contain a maximum of ten columns. By default, eight columns are included. To add three spectral matching columns, one other column must be removed. f) After the run is complete, view the Analysis Report (either from the Results or the Reports menu). Matches will be reported. If there was no match, the cell in the Sample Table will be empty Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

59 View Spectral Data for Another Sample To view the 3D plot, chromatogram plot, and spectrum plot for another sample: 1 Close the Spectral Data window by clicking the x in the upper right corner. The Run Results window appears. 2 On the Run Results window: a) Select a different sample from the Samples drop-down list. b) Optionally, select a different data channel and/or analysis. 3 Click the Spectral Data button to view the chromatogram and spectral plots for the selected sample. 4 Repeat for any other samples. View Results and Spectral Data Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-47

60 Fraction Collection Simulation Fraction Collection Simulation How to Simulate Fraction Collection by Slope or Level Optimize fraction collection settings using the fraction collection simulator: 1 Create a method that includes fraction collection by slope or level. 2 Browse in a representative example chromatogram from a previous run s results. a) Click the Application Run Results tool ( ). b) From the drop-down menu, select the application that was run to obtain the results that will be overlaid and the channel for which to overlay the results. c) Double-click the Run Name/Method/Channel. The Select Iteration dialog appears. d) On the Select Iteration dialog, do either of the following: Double-click the iteration number. Click to select the iteration number and then click OK. The chromatogram overlays in the method control beginning at the Start Data Collection task. Optionally, modify the color or remove the chromatogram trace. For more information, refer to How to Modify the Appearance of Overlaid Results. If using the Conditional Fraction Collection task with more than one data channel, complete steps a d for each channel. The fraction start and fraction end lines overlay the chromatogram showing the current fraction collection settings in effect. If variables were used for any of the fraction collection parameters, modify the Default Value for the variable in the Variable Properties dialog. If the Value is not modified, 0 (the default Value) will be used in the simulation. 3 Optionally, modify the fraction collection parameters. 4 Click the Update Chromatogram tool ( ). The fraction start and fraction end lines update based on the modifications to the fraction collection parameters Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

61 How to Modify the Appearance of Overlaid Results Click to open the Results List dialog. The Results List dialog in the Method Builder provides previews of overlaid results for all channels and allows for changing the color of the data trace of each overlaid result or removing overlaid results. To change the color of a trace, select the check box next to the preview of the trace, and then click Color. Select a color from the color palette. To remove overlaid results, select the check box next to the preview of the trace, click Remove, and then click OK. Fraction Collection Simulation Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-49

62 View Diagnostics View Diagnostics To check the status of the Flexar SQ 300 MS Detector, view the diagnostics. To generate a diagnostics report: 1 In the Project Library, select an application containing a method using a configuration that includes a Flexar SQ 300 MS Detector and then click Run. The Application Run window appears. 2 Under Method Configuration, select a method using a configuration that includes a Flexar SQ 300 MS Detector (if not already selected), and then click. 3 Click to access the Manual Control window. 4 Issue the Mass Spec Diagnostics command to the MS Detector: a) From the Task/Command List, drag and then drop the Mass Spec Diagnostics command into the workspace. b) Set the properties. 1) For the Instrument Name property, specify the name assigned to the Flexar SQ 300 MS Detector in the configuration (by default it is MS Detector). 2) For the Synchronize property, the default selection of On should be used. 3) For the Auto Save property, select TRUE to save or FALSE to not save the diagnostics report on the Tune/Diagnostics tab in the Results. c) Click Run. The diagnostics report appears. 5 When viewing the diagnostics, there are options for viewing, saving, or printing the report. Saved diagnostic reports are saved to a user-specified location as.html files. It is recommended to save check tune results in a folder for tracking of long-term trends. Select one of the following forms of the diagnostics report to view: Short Diagnostics - Displays run-critical diagnostics. All Current Values - Displays the actual values at the time requested for all diagnostic parameters. All Diagnostics - Displays values entered and actual values for all diagnostic parameters. 6 To view the diagnostics as recorded at the beginning of a sample for which data was collected in the application run: a) Open the Results window - Results tab. b) Right-click on the run name and then select View Diagnostics. c) Double-click the sample to view the diagnostics as recorded at the beginning of that sample. d) Select a form of the diagnostics report to view: Short Diagnostics, All Current Values, or All Diagnostics. e) Optionally, print or save the report Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

63 View Status While in the Application Run window, view the status of the Flexar SQ 300 MS Detector by hovering over the status box and viewing the tooltip information. This information is not updated while collecting data. View Diagnostics Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-51

64 Manage Maintenance Alarms Manage Maintenance Alarms A feature of the software is the ability to set one or more maintenance alarms. The default maintenance alarm (which can be edited) is a reminder to auto tune the Flexar SQ 300 MS Detector. Depending on the application and frequency of use of the Flexar SQ 300 MS Detector, set the alarm to remind hourly, daily, weekly, monthly, or yearly. The default setting is off. Edit this alarm or add additional alarms from the Administrative Tools menu by selecting the Maintenance Alarms option. Create an Alarm To create a custom alarm: 1 Click Add. 2 When the Alarm Name dialog appears, type a name and then click OK. 3 Select a Start day and time, the recurrence (Period), and optionally, type a message. 4 By default, any new alarms are turned On. Optionally, select Off to turn off an alarm temporarily or permanently. 5 Click Save. Modify an Alarm To modify an existing alarm: 1 Make desired changes to the alarm settings. 2 Click Save. Rename an Alarm To rename an alarm: 1 Click Rename. 2 Type the new name for the alarm. 3 Click OK. 4 Click Save Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

65 Alarm Reminder A clock will be visible when accessing the Application Run window if an alarm is due soon. Hover the mouse over it or click to view a listing of the alarms due soon. If an alarm is due or overdue, a yellow bar appears in the Application Run window informing that Maintenance Alarms need attention. Click here. Click the bar, select an alarm, and then click Done or Remind Me Later. If Remind Me Later was selected, the yellow bar will appear again the next day. Manage Maintenance Alarms Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 2-53

66 Maintenance 3 This chapter describes the routine maintenance procedures required to keep the Flexar SQ 300 MS Detector in proper working condition and to ensure the highest possible level of performance. It includes maintenance checks that should be done on a daily basis and maintenance procedures that should be done periodically depending on use. You should perform only the maintenance procedures described in this chapter. If additional maintenance is required, contact your local Gilson representative. To reduce the chance of electrical shock, do not remove covers that require tool access. No user serviceable parts are inside. Before using any cleaning or decontamination methods except those specified by the manufacturer, check with your local Gilson representative that the proposed method will not damage the equipment. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 3-1

67 Maintenance 3 Periodic Maintenance Periodic Maintenance The frequency of maintenance required will be determined by the amount of use the system receives, the cleanliness of the environment, and the number and nature of the samples being purified. Periodic ESI Source Cleaning On a periodic basis clean: the probe plate around the source the end plate and end plate cap the capillary entrance Keeping the probe plate, the source area, the end plate cap, and capillary entrance clean will keep sensitivity high and maintain a steady signal in the Flexar SQ 300 MS Detector. To prevent contamination, always wear powder-free gloves when handling the source. Never block off the capillary entrance, as this will cause a backup of oil into the vacuum chamber and can damage the Flexar SQ 300 MS Detector. Never spray methanol or any solvent directly into the capillary entrance, as this can damage the Flexar SQ 300 MS Detector. 1 Run the Mass Spec Single Quad Start Up task (in Manual Control). Change the Dry Gas Flow to 2000 ml/min. Change the Dry Gas Temperature to 50 C. Change the Nebulizer Gas Pressure to 0 psi. It will take about 30 to 40 minutes for the instrument to cool down enough to handle. 3-2 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

68 Maintenance 3 2 Carefully remove the probe from the probe plate. Take care not to damage the probe tip when removing the probe from the probe plate. Holding on to probe Press in button on the probe plate Periodic Maintenance Remove probe Press button in all the way on the probe plate 3 Loosen the finger-tight screw securing the source door. 4 Open the source door. Loosen this screw Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 3-3

69 Maintenance 3 Periodic Maintenance 5 Spray a lint-free wipe with methanol and then thoroughly wipe the inside of the probe plate. 6 Spray methanol on a lint-free wipe and then thoroughly clean the inside of the source. 3-4 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

70 Maintenance 3 7 Remove the end plate cap and clean the front surface of the end plate. 8 Clean the end plate cap. end plate cap Periodic Maintenance 9 Lightly dampen a lint-free wipe with a small amount of methanol and then gently clean the front of the capillary entrance. 10 Replace the end plate cap. 11 Close the source door and then secure the door shut with the finger-tight screw. 12 Reinstall the probe. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 3-5

71 Maintenance 3 Replace the Probe Needle Replace the Probe Needle When adjusting the probe needle position, the spray should be a fine mist that is steady (not pulsing) and is symmetrical. If it is not, the probe needs to be cleaned, replaced, or adjusted. 1 Turn the adjustment locking nut clockwise to allow it Phillips set screw adjustment locking nut to move freely. 2 Unscrew the probe needle depth adjustment nut counterclockwise until it comes off. 3 Remove the Phillips set screw by turning counterclockwise. probe needle depth adjustment nut Note: The keyed metal insert is spring-loaded. Secure the insert while removing the set screw. keyed metal insert 3-6 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

72 Maintenance 3 4 With the screw removed, remove the keyed metal insert. The probe, the insert with needle attached, and the Phillips set screw with rubber O-ring are shown at right. Replace the Probe Needle 5 To remove the old probe needle, use a ¼ inch wrench and turn the fastening nut counterclockwise to loosen. 6 Take the new probe needle and place it into the wider side of the ferrule. Use the wrench to loosen the fastening nut on the probe needle 7 Thread the probe needle through the ferrule and fastening nut. The probe needle through ferrule and fastening nut Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 3-7

73 Maintenance 3 Replace the Probe Needle 8 Push the needle flat against the rubber surface of the ferrule. This creates a stopping point for the needle and will prevent dead volume. Push the needle flat against the ferrule 9 Use a finger tight nut to screw in the red or black PEEK tubing into one side of the keyed metal insert. Red PEEK tubing into one end of keyed metal insert 10 At the other side where the probe needle is flush, screw the probe needle into the keyed metal insert until it becomes just snug. Screwing in the probe needle 11 To prevent dead volume, gently pull the probe needle out slightly then push the probe needle in until the needle stops against the back side of the PEEK tubing on the other side. 12 Finger-tighten the fastening nut. Use a ¼ inch wrench to turn the fastening nut 1/8 turn clockwise to complete the tightening. 13 Verify that the probe needle is snug. 14 Remove the PEEK tubing and finger tight nut. Tightening the fastening nut 3-8 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

74 Maintenance 3 15 Push the probe needle into the small hole at the base of the probe. Gently twist and turn the probe needle until it falls through the hole. 16 Place the slot into the groove on the probe. 17 Press the keyed metal insert down with your finger and check to see that the probe needle goes in and out. 18 Reinstall the Phillips set screw in the probe. Replace the Probe Needle Pushing the probe needle into the hole 19 Screw in the probe needle depth adjustment nut until the needle pops out 1/8 inch. 20 Turn the adjustment locking nut counterclockwise to lock in the probe needle depth adjustment nut. Probe needle is out 1/8 inch Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide 3-9

75 Maintenance 3 Maintenance of the Roughing Pump Maintenance of the Roughing Pump You should never service the roughing pump or oil mist filter. Refer to the documentation supplied with pump and oil mist filter for additional information. Gas Ballasting the Roughing Pump Once a week you should gas ballast the roughing pump, otherwise oil will build up in the oil filter chamber, by doing the following: 1 Slowly open the gas ballast by turning the knob counterclockwise. 2 The oil will drain through the oil return. gas ballast knob oil level before the oil has been drained 3 When the oil level is low, turn the gas ballast knob clockwise to close. oil level after the oil has been drained 3-10 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

76 Splitter Connections and Control A The Rheodyne MRA splitter is controlled by contact signals triggered by TRILUTION LC. Note: To control the splitter as a GEARS device, instead of by contact control, use the GEARS MRA Splitter Control (part number ). Refer to the documentation supplied with it for more information. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide A-1

77 Splitter Connections and Control A Electrical Connections When making the contact connection: Use contacts 4 and GROUND on the splitter Use output 2 on the GX-271 Prep or 215 Liquid Handler Use output 3 on the GX-281 Prep Liquid Handler Make Connections To make connections, the following are needed: 2-conductor cable (22 30 gauge for each wire) wire insulation stripper small-blade screwdriver To make connections with the 2-conductor cable: 1 Cut the cable into pieces of appropriate length. 2 Strip about 3 mm of insulation from each end of the cable. 3 Remove the terminal block connector from the liquid handler. Insert each wire into the appropriate terminal on the terminal block connector. Note: When making connections, be sure to maintain the correct orientation of the connector relative to the port. Push the wire all the way in; then tighten its corresponding pin screw. 4 Reconnect the terminal block connector to the liquid handler. Push the connector in as far as it will go. It is designed to fit snugly into its receptacle. 5 Connect the opposite ends of the wires to the other device(s). Be sure to match ground connections. 6 Label each cable to identify the purpose of the connection. Splitter Connections and Control Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide A-2

78 Splitter Connections and Control A Control Although the splitter is controlled by contact signals triggered by TRILUTION LC, some setup of the splitter is required. 1 Set the splitter for local control by pressing the LOCAL REMOTE button on the splitter until the light next to LOCAL illuminates. 2 Refer to the following table to determine the split factor, based on the flow rate. Keep this information available for reference. Split Ratio HPLC Flow Rate (ml/min) Splitter Connections and Control Source: Data from MRA Operating Manual, Rheodyne, L.P., A(1), 8/01. 3 Set the split factor: a) Press the SET button. The display will flash. b) Press the arrow button until the desired split factor is displayed. c) Press the SET button again. Two horizontal lines will appear on the display, indicating that the split factor has been set. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide A-3

79 Source Change B The Flexar SQ 300 MS Detector is shipped with the ESI source installed. Refer to the instructions in this appendix to remove the ESI source and install the atmospheric pressure chemical ionization (APCI) source (ordered separately, part number ). Also refer to these instructions when re-installing the ESI source before running an auto tune or check tune. Refer to the appropriate instructions: Replace ESI Source with APCI Source Replace APCI Source with ESI Source Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide B-1

80 Source Change B Source Change Replace ESI Source with APCI Source Remove the ESI Source The operating temperature of the source is often 350 C. Allow minutes for it to cool before handling it. To prevent contamination, always wear powder-free gloves when handling the source. To remove the ESI source: 1 Start TRILUTION LC. 2 In the Project Library, select an application containing a method using a configuration that includes a Flexar SQ 300 MS Detector and then click Run. The Application Run window appears. 3 Under Method Configuration, select a method using a configuration that includes a Flexar SQ 300 MS Detector (if not already selected), and then click. 4 Click to access the Manual Control window. 5 Run the Mass Spec Single Quad Standby task (in Manual Control). B-2 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

81 Source Change B 6 Disconnect the tubing from the MS source probe. Source Change 7 Loosen the finger-tight screw securing the source door and then open the source door. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide B-3

82 Source Change B Source Change 8 Lift the ESI source door straight up and off of the hinges. 9 Place the source on a clean surface in a safe place for future use. B-4 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

83 Source Change B Install the APCI Source To install the APCI source: 1 Align the pins on the APCI source with the hinges on the Flexar SQ 300 MS Detector. Source Change 2 Slide the APCI source down until the pins are in the hinge and the source can rotate freely on the hinge. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide B-5

84 Source Change B Source Change 3 Close the source door and then tighten the black knob on the front of the source. 4 Plug the cable from the APCI source into the port (labeled APCI PROBE) on the front panel of the Flexar SQ 300 MS Detector, and then twist it until secure. B-6 Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide

85 Source Change B 5 Connect tubing from port 1 on the diverter valve to the MS source probe: a) On one end of the PEEK tubing (labeled A, part number ), use the fingertight fitting (part number ) to connect to the MS source probe. b) Use the PEEK nut and ferrule (part number ) to connect the other end of the tubing to port 1 on the diverter valve. For more information, refer to the plumbing diagram (part number LT309139). Source Change 6 Start TRILUTION LC. 7 In the Project Library, select an application containing a method using a configuration that includes a Flexar SQ 300 MS Detector, and then click Run. The Application Run window appears. 8 Under Method Configuration, select a method using a configuration that includes a Flexar SQ 300 MS Detector (if not already selected), and then click. 9 Click to access the Manual Control window. 10 Run the Mass Spec Single Quad Start Up task (in Manual Control). Select the Source Type: APCI. The default values are acceptable. Wait 5 minutes before operating for running samples. This allows the Flexar SQ 300 MS Detector to reach proper operating temperature. Gilson LC/MS Purification System in TRILUTION LC Software v3.0 User s Guide B-7