MassLynx NT Guide to Data Acquisition

Transcription

1 MassLyx NT Guide to Data Acquisitio Versio th August 2000 Page i

2 MassLyx NT Guide to Data Acquisitio The software described i this book is furished uder a licece agreemet ad may be used oly i accordace with the terms of that agreemet. Copyright Notice Micromass Ltd believes that the iformatio i this publicatio is accurate. However the iformatio is subject to chage without otice ad should ot be costrued as a cotractual udertakig by Micromass Ltd. Despite the care which has bee give to the preparatio of this publicatio, Micromass Ltd accepts o resposibility for ay loss or ay other matter which may arise from ay error or iaccuracy which may iadvertetly have bee icluded. Copyright (C) Micromass Ltd. All Rights Reserved. No part of this publicatio may be copied without the express writte permissio of Micromass Ltd. Trademarks Micromass is a registered trade mark of Micromass Limited (Reg. U.S. Pat. & Tm. Off.). MassLyx is a registered trademark of Micromass Ltd. Widows is a trademark of Microsoft Corporatio. Other product ames metioed i this maual may be trademarks or registered trademarks of their respective compaies ad are hereby ackowledged. Page ii

3 Table of Cotets Acquirig Data a Overview 1 Acquirig Data a Overview 1 Gettig Started the basic steps for acquirig data 1 The Acquisitio Cotrol Pael 5 Itroductio 5 Cofigurig the Ilet System 6 Settig Istrumet Data Thresholds 7 Chagig Lab ad User Iformatio 17 Other Meu Items 17 Istrumet Tuig 21 Itroductio 21 The Tue Page 21 The Tue Page Toolbar 23 Chagig Tue Parameter Settigs 24 Pritig Tue Iformatio 24 Experimetal Record 24 Savig ad Restorig Parameter Settigs 25 Modifyig the Peak Display 27 AutoTue 28 Scope Parameters 31 Chagig Ilet Heaters 32 Gas Cotrols 33 Ramp Cotrols 34 Vacuum 35 Resettig the Zero Level 35 Cotrollig Readbacks 36 Startig a Acquisitio from the Tue Page 37 Q-Tof Tuig 39 Mass Calibratio 45 Mass Calibratio 45 How a calibratio is formed 45 Calibratio Types 45 Overview of the calibratio process 46 The Calibratio Dialog 47 Parameters which cotrol Calibratio 48 QTOF Accurate Mass 53 The Calibratio Report 54 Other Calibratio Facilities 55 Editig a Referece File 57 Savig ad Restorig Calibratios 57 Page iii

4 The Fuctio List Editor 61 Itroductio 61 Addig a New Fuctio 62 Modifyig a Existig Fuctio 62 Removig a Fuctio 62 Chagig a Fuctio s Start ad Ed Times 63 Settig a Solvet Delay 63 Aalog Chaels 63 Savig ad Restorig a Fuctio List 64 Settig up a Full Sca Fuctio 66 Settig up a SIR Fuctio 69 Settig up a MS/MS Fuctio 70 Settig up a MRM Fuctio 74 Q-Tof Fuctio Editig 75 Survey Fuctios 75 Automatic Fuctio Switchig 78 Acquirig Data 93 Startig a Acquisitio 93 Sigle Sample 93 Multiple Samples 94 Moitorig a Acquisitio 97 The Acquisitio Status Widow 97 The MCA Display 97 Chromatogram Real-time Update 98 Spectrum Real-time Update 98 Stoppig a Acquisitio 99 Automatic Startup ad Shutdow 100 The Shutdow Editor 100 The Auto Cotrol Tasks Page 101 The Shutdow Editor Toolbar 103 Savig/Loadig Startup ad Shutdow Files 104 Pritig Startup ad Shutdow Files 105 Creatig Startup ad Shutdow Files 105 Ruig Startup ad Shutdow Files 105 Idex 109 Page iv

5 Acquirig Data a Overview Acquirig Data a Overview Chapter 1 This chapter provides iformatio o usig the acquisitio system. It will explai how MassLyx is used to tur o ad tue the mass spectrometer, how the istrumet is the calibrated ad the fially how you ca acquire data oto the computer s hard disk. Gettig Started the basic steps for acquirig data This sectio gives a overview of the basic steps required to do a acquisitio ad the order i which these steps should be performed. Each of the steps ivolved i acquirig data is described i much greater detail i its ow sectio later i this chapter. 1. Load MassLyx Load MassLyx by double clickig o the MassLyx ico i the MassLyx group. If MassLyx Security is eabled the MassLyx Logi dialog will be displayed. Eter your Logo Name ad Password. MassLyx will be loaded ad commuicatios with the istrumet will be iitialised. 2. Cofigure the Ilet System If ecessary choose Select Iterface from the Acquisitio Cotrol Pael Cofigure meu ad select which type of ilet system you are usig. Next choose Ilet from the Acquisitio Cotrol Pael Methods meu. This allows you to set up the ilet parameters for your chose ilet system e.g., the GC temperature ramp if you are usig a GC system. 3. Prepare istrumet for use The istrumet should be prepared for use as described i the Users Guide supplied with your istrumet, if it is i a veted state it should be pumped dow usig the vacuum cotrols. The istrumet should the be tured ito operate. 4. Tue the istrumet The istrumet must be tued to obtai the best coditios for ioizatio. Agai this should be doe i accordace with the istructios give i the Users Guide supplied with your istrumet. Page 1

6 5. Calibrate the mass scale The mass scale of the istrumet must be correctly calibrated so that the masses reported for acquired data are correct. Calibratio of the mass scale ivolves itroducig a referece compoud ito the system, acquirig some data ad the comparig the result with the expected masses for the referece compoud. A calibratio curve is produced ad used to correct for ay errors i the mass scale. 6. Acquire some data You ca ow acquire some data, either for a sigle sample or for a umber of samples, itroduced ito the system usig a autosampler. To do this you must set up a method for scaig the system (referred to as a fuctio) ad if you are usig a programmable ilet system, you must also set up ay ilet related parameters, icludig those that cotrol a autosampler if fitted. You ca the start the acquisitio ad let it ru for a programmed duratio or stop it maually before that time. 7. Moitorig data acquisitio The progress of the acquisitio ca be moitored either by selectig the Status display which gives a sca by sca report of acquisitio statistics, or by viewig the data that is beig acquired i real time usig the Spectrum or Chromatogram displays. Page 2

7 Notes Page 3

8 Notes Page 4

9 The Acquisitio Cotrol Pael Itroductio Chapter 2 All MassLyx Acquisitio fuctios are accessed from, ad maaged by, the Acquisitio Cotrol Pael. I order to perform a acquisitio you must first tell the system about the istrumet cofiguratio that you wish to use. The Acquisitio Cotrol Pael will the cofigure itself to allow you access to the cotrols that are appropriate to the iteded task. MassLyx ca be cofigured so that the Acquisitio Cotrol Pael is automatically displayed whe MassLyx is ru, otherwise the Acquisitio Cotrol Pael is displayed whe you select Cotrol Pael from the MassLyx Ru meu or press the toolbar butto. The picture displayed will reflect your curret Istrumet ad Ilet cofiguratio. Figure 2.1 shows the Cotrol Pael display for a Quattro II with a GC ad Autoijector. Acquisitio cotrols ca be accessed either with the meu bar at the top of the Cotrol Pael, or i the case of the mass spectrometer tue page or the ilet editors, by double clickig o the relevat part of the Cotrol Pael picture. To automatically load the Acquisitio Cotrol Pael with MassLyx To automatically load the Acquisitio Cotrol Pael with MassLyx select Autoload from the Acquisitio Cotrol Pael Cofigure meu. A tick will appear ext to this meu optio whe it is selected. To select the Tue widow from the Cotrol Pael Double click o the mass spectrometer part of the istrumet picture -or- Choose Tue Mass Spectrometer from the Acquisitio Cotrol Pael Istrumet meu. -or- Press CTRL-T Figure 2.1 The Acquisitio Cotrol Pael Page 5

10 Cofigurig the Ilet System Before you use the Acquisitio Cotrol Pael you must first cofigure it to support the ilet system fitted. This is doe usig the Select Iterface dialog box. MassLyx will display a list of the ilet systems which you selected durig istallatio of the MassLyx software. If you add additioal ilet systems at a later date you may eed to reistall MassLyx to add the cotrol software for the ew ilet system. Oce you have cofigured the ilet system, MassLyx will give you access to the parts of the acquisitio system that are appropriate to the ilet selected ad the Tue page will chage if the ilet selected requires a chage of ioizatio mode. Figure 2.2 Select Ilet Dialog The list of ilet optios, which appears i the Select Iterface dialog, reflects the ilet systems which were selected whe MassLyx was istalled. If, at a later date you add a ew ilet system or chage oe of your existig ilets you may eed to reistall MassLyx to gai access to the cotrol software for the ew ilet system. To chage the ilet system 1. Choose Select Iterface from the Acquisitio Cotrol Pael Cofigure meu. 2. Scroll dow to the required ilet system. 3. Select the ilet. 4. Press the OK butto. The Cotrol Pael will ow re-cofigure itself to show the ew ilet system. Page 6

11 Settig Istrumet Data Thresholds MassLyx has several parameters that allow you to cotrol how the trasputer system pre-processes data before it is set to the host computer. These parameters are cotaied i the Istrumet Data Thresholdig dialog. Istrumet Data Thresholdig allows you to specify what type of data you wish to acquire ad write to disk ad what type of data you wish to discard ad ot write to disk. Limitig the amout data stored o disk ca be particularly desirable whe acquirig cotiuum data ad doig log LC rus. To chage Data Thresholdig 1. Choose Set Istrumet Threshold from the Acquisitio Cotrol Pael Istrumet meu. 2. Make required chages to the iformatio. 3. Press the OK butto. The data thresholdig dialog displayed will deped o the type of istrumet beig used. For Quattro I, Bio-Q, Trio-2000 ad istrumets the followig dialog will be displayed. Figure 2.3 Istrumet Data Thresholdig Dialog (Quattro I, Bio-Q, Trio-2000 ad ) Page 7

12 For Quattro II ad Platform istrumets the followig dialog will be displayed. Figure 2.4 Istrumet Data Thresholdig Dialog (Quattro II ad Platform) Pressig the Advaced butto i this dialog will display the Advaced Thresholdig Parameters dialog. Figure 2.5 Advaced Istrumet Data Thresholdig Dialog (Quattro II ad Platform) Pressig the OK butto stores ay chages made i the Istrumet Data Thresholdig dialog ad i the Advaced Istrumet Data Thresholdig dialog if preset. For Quattro II ad Platform II istrumets the tue page will be updated immediately if it is ope, i.e. the effect of Baselie Level ad Io Coutig chages are show o the active tue segmets. For other istrumets the ew parameters will be dowloaded at the start of the ext acquisitio sca. Page 8

13 Pressig Cacel discards ay chages made ad closes the Istrumet Data Thresholdig dialog. Pressig the Default butto loads default values for all parameters ad switches off all the istrumet data thresholdig features. MaxEt Users Profile Data Note: The MaxEt algorithm eeds to accurately measure oise withi a data file. For this reaso the Io Coutig Threshold should be set to zero whe acquirig data which will be aalysed usig MaxEt. The cotrols for profile data allow you to cotrol the amout of data that is collected durig a cotiuum data acquisitio. By default MassLyx will collect oe data poit every 8th of a dalto. If for istace you were scaig your istrumet from mass 50 to mass 1300 you would collect ad save data poits per sca, ad as each poit requires 6 bytes of disk space, every sca would take 1/17 of a megabyte of disk space. Obviously if you use this type of scaig i cojuctio with chromatography, the data file sizes will grow to be eormous. I additio to the disk space issue, collectig this amout of data puts a heavy demad o the trasputer system, which will ultimately effect the sca rates at which you ca collect data. The ability to use a threshold with cotiuum data is therefore highly desirable as it allows you to disregard data that is oise yet save the complete peak profile for the real data, thus retaiig the iformatio i the data while reducig disk space requiremets. To use this facility you should set Data Storage to Compressed. Havig selected Compressed mode, Io Coutig Threshold is used to set the itesity level below which a data poit will be igored. This feature is discussed later. Data Storage Whe Data Storage is set to Stadard every mass/itesity pair is stored to file, regardless of their values, i.e. zero itesities. This mode must be used for acquisitios with MCA fuctios Whe Data Storage is set to Compressed zero itesity values are ot stored to file so this produces smaller file sizes. Compressed Data Storage mode caot be used for MCA acquisitios. Baselie Level Is used to lift/drop the baselie to see more/less of the oise. It is used whe Io Coutig Thresholdig is ot eabled (i.e. set to zero), to set the positio above zero of the baselie. The baselie level would typically be set to a value of 1. If you wish the baselie to appear higher the you ca icrease the value of the baselie level parameter. It is possible to use a egative baselie. This reduces the oise see ad acts as a form of thresholdig to be applied to 1/16th amu type samples. This will take place after io coutig ad therefore have a less sigificat effect. If you wat to see more oise use a positive value. Do ot use a positive value for the Baselie Level if you are usig Io Coutig Thresholdig or Compressed Data Storage mode. Page 9

14 The Poits per dalto parameter ca have oe of three values, 4, 8 or 16. Selectig 8 poits will allow you to acquire data twice as quickly as selectig 16, ad will result i data files which are approximately half as big as those acquired at 16 poits per dalto. Selectig 4 poits will allow you to acquire data twice as quickly as selectig 8, ad will result i data files which are approximately half as big as those acquired at 8 poits per dalto. Acquirig data at 16 poits per dalto gives the greatest possible resolutio. Acquirig data at 4 poits per dalto gives data with a smoothed appearace. Cetroid Data SIR Data For cetroided data, there are three parameters that you ca set. Dyamic Rage Cotrols the size (i terms of bytes) of the umber used to store a acquired itesity value ad also the precisio with which a mass umber is stored. The stadard dyamic rage settig allows the itesity values to be stored usig 4 bytes ad masses to be stored to a 1/128 amu precisio. The high dyamic rage is seldom used but allows 5 bytes for itesity ad 1/1024 amu mass precisio. Data Threshold Sets a itesity level below which peaks detected will be igored. This is useful if you wish to cocetrate o larger peaks of iterest ad cut dow o the size of acquired data files. Peak Acceptace Allows you to igore the miimum peak area calculated durig the presca thus allowig smaller peaks through. Settig Peak Acceptace to high activates this ad will result i more peaks beig stored per sca. The default value is low. The SIR Baselie Level is used whe Io Coutig Thresholdig is ot eabled (i.e. set to zero). It is used to set the positio above zero of the baselie. The baselie level would be typically be set to a value of 1. If you wish the baselie to appear higher the you ca icrease the value of the baselie level parameter. Io Coutig Thresholdig Note: For Quattro II ad Platform istrumets these parameters are accessed by pressig the Advaced butto i the Istrumet Data Thresholdig dialog. Io Coutig Threshold is used to set the itesity level below which a data poit will be igored. The Io Coutig Threshold ca be set to values betwee 0 ad 25, the higher the umber you eter the more data will be discarded. Note: To disable the Io Coutig Threshold set the value to zero. If you wish to use the Io Coutig Threshold facility a value of 6 is suitable for most data. This threshold is applied to all acquisitios, regardless of scaig mode. It is also the most sigificat of all of the data maipulatio variables because it is the oe applied first to the raw data. Page 10

15 Whe a acquisitio is started the istrumet performs a "presca" with the io beam switched off so that the electroic oise level of the acquisitio system ad its stadard deviatio ca be measured. The Io Coutig Threshold level that you eter is multiplied by the stadard deviatio of the oise to determie the itesity level to be used. If you set a value of zero the itesity level will be set so that it sits right i the middle of the oise which would mea that roughly half of the oise data would be acquired. If you set a value of 20 the threshold would sit well above the oise level, so very little oise data would be acquired. Coversely a value of 1 would place the threshold oly just above the oise so almost all of the data would be acquired. Importat Note: Whe usig a io coutig threshold you should set the profile data baselie level ad SIR data SIR baselie level to zero. Figure 2.6 to Figure 2.7 shows the effects of chagig the Baselie Level ad Io Coutig Threshold parameters. A series of acquisitios were doe usig Heptacosa ad acquirig cotiuum data. Each data file cotais 18 scas. The data file AB0000 is ot thresholded, both Baselie Level ad Io Cout Threshold were set to zero. Figure 2.6 shows the effect of icreasig the Baselie Level parameter. Baselie = 0, Io Coutig = 0 AB (0.128) Sm (SG, 2x1.00); Cm (1:18) % Baselie Level=5 Sca EI+ 6.61e4 0 AB (0.128) Sm (SG, 2x1.00); Cm (1:18) % AB (0.232) Sm (SG, 2x1.00); Cm (1:18) % AB (0.128) Sm (SG, 2x1.00); Cm (1:18) 100 % Baselie Level=2 Baselie Level=1 Baselie Level=0 Sca EI+ 6.03e4 Sca EI+ 6.19e4 Sca EI+ 4.48e Da/e Figure 2.6 Effect of icreasig Baselie Level o Heptacosa spectrum Page 11

16 Figure 2.7 shows the effect of icreasig the Io Coutig threshold o a part of the spectrum, which cotais oly backgroud oise. The bottom trace was acquired with the Io Coutig Threshold set to zero, for subsequet traces the Io Coutig Threshold was set to 1, 2, 4, 6 ad 25. As the Io Coutig Threshold is icreased the amout of oise stored is reduced, the ormalizig itesity value at the top right had corer of the trace is also reduced. Baselie = 0, Io Coutig = 25, Data Storage = Compressed AB (0.128) Cm (1:18) 100 Sca EI+ 1 % 0 AB (0.128) Cm (1:18) Sca EI+ 676 % AB (0.128) Cm (1:18) % Sca EI Da/e Baselie = 0, Io Coutig = 0 AB (0.232) Cm (1:18) Sca EI e % 0 AB (0.128) Cm (1:18) 100 % Sca EI+ 1.42e AB (0.128) Cm (1:18) % Sca EI e3 0 Da/e Figure 2.7 Effect of icreasig Io Coutig Threshold o oise i Heptacosa spectrum The value of the Io Coutig Threshold should be set such that backgroud oise is removed without sigificatly reducig the itesity of the smallest peaks which are of iterest. Page 12

17 Figure 2.8 shows the effect of icreasig the Io Coutig threshold o a part of the spectrum, which cotais a low itesity peak. As the Io Coutig Threshold is icreased beyod a certai value the peak becomes arrower ad its itesity is reduced as the thresholdig rejects part of the geuie sigal. I this case a Io Coutig Threshold value of 4 would be suitable. Baselie = 0, Io Coutig = 4, Data Storage = Compressed AB (0.128) Sm (SG, 2x1.00); Cm (1:18) 100 Sca EI+ 1 % Io Coutig Threshold=25 0 AB (0.128) Sm (SG, 2x1.00); Cm (1:18) Sca EI+ 1.53e4 % Io Coutig Threshold=6 0 AB (0.128) Sm (SG, 2x1.00); Cm (1:18) Sca EI+ 2.84e4 % Io Coutig Threshold=4 0 Da/e Baselie = 0, Io Coutig = 2, Data Storage = Compressed AB (0.232) Sm (SG, 2x1.00); Cm (1:18) Sca EI e4 % Io Coutig Threshold=2 0 AB (0.128) Sm (SG, 2x1.00); Cm (1:18) Sca EI+ 4.30e4 % Io Coutig Threshold=1 0 AB (0.128) Sm (SG, 2x1.00); Cm (1:18) Sca EI+ 4.48e4 % Io Coutig Threshold=0 0 Da/e Figure 2.8 Effect of icreasig Io Coutig Threshold o low itesity peak i Heptacosa spectrum Page 13

18 Table 2.1 gives a summary of the parameters used to acquire each of the data files ad shows the % reductio i file size which resulted from usig the series of Io Coutig Thresholds ad storig data i Compressed format. File Name Baselie Level Io Cout Threshold Data Storage.DAT File Size (Bytes) % savig o AB0000 AB Stadard 1,102,926 AB Stadard 0 AB Stadard 0 AB Stadard 0 AB Compressed 1,014,942 8 AB Compressed 998, AB Compressed 432, AB Compressed 347, AB Compressed 185, Table 2.1 Summary of Istrumet Data Thresholdig parameters used to acquire files AB0000 to AB0250. Multiplet Threshold Note: For Quattro II ad Platform istrumets these parameters are accessed by pressig the Advaced butto i the Istrumet Data Thresholdig dialog. The real-time peak detectio algorithm i the acquisitio system uses a value of peak separatio called the Multiplet Threshold to determie whether two peaks will be detected as two separate peaks or combied ito oe peak. If two peaks are separated by less tha Multiplet Threshold the they will be combied ito a sigle peak. However the value of the multiplet threshold eeds to icrease as the mass scale icreases sice peaks will ted to broade at higher masses. For this reaso a ramp may eed to be applied to the value of the multiplet threshold so that it icreases with the mass rage. This is doe usig the parameters Ramp Start ad Fial multiplet threshold. Page 14

19 Advaced Istrumet Data Thresholdig parameters (Quattro II ad Platform istrumets oly) These parameters are accessed by pressig the Advaced butto i the Istrumet Data Thresholdig dialog. Figure 2.9 Advaced Istrumet Data Thresholdig Dialog (Quattro II ad Platform) Profile Data Spike Removal Spikes are distiguished from real data by the fact that spikes are very arrow ad also very itese whe compared to their immediate eighbors. Data poits that are determied to be spikes are removed by settig the value of this data poit to the average of its immediate eighbors. Note that the use of Spike Removal does ivolve some additioal processig while acquirig. For this reaso the use of Spike removal will reduce the maximum achievable acquisitio rates by approximately 30%. Use Spike Removal Select this box to perform spike removal durig a acquisitio. (Note this will ot be reflected i the tue page). Miimum Spike Itesity Itesity threshold below which spikes will be igored. Take this value from the Tue page itesities. A very low itesity sigal may iclude sigle io evets that ca be combied to produce sigificat peaks. For this type of data the Miimum Spike Itesity should be set to a suitable value such that these sigle io evets are ot discarded as spikes. Spike Percetage Ratio Ratio used to determie if a data poit is a spike by comparig the data poit to its immediate eighbors. If the Spike Percetage Ratio is set to 30%, the if at 30% of its full itesity the data poit is still more itese tha both its immediate eighbors it will be regarded as a spike. To express this as a ratio, the maximum allowed itesity ratio betwee a data poit ad its immediate eighbors is 3:1. Spike Percetage Ratio set to 50% is equivalet to a ratio of 2:1. Spike Percetage Ratio set to 20% is equivalet to a ratio of 5:1. Page 15

20 Profile Data Ehaced TIC Calculatio Eable TIC Ehacemet Select this box to produce a ehaced TIC for the acquisitio. Note that the use of TIC Ehacemet does ivolve some additioal processig while acquirig. For this reaso the use of TIC Ehacemet will somewhat reduce the maximum achievable acquisitio rates. TIC summatio type The TIC ca be either the sum or BPI of those itesities withi the specified rages. Itesity Acceptace Parameters These parameters specify a miimum ad maximum itesity ad a mass rage for the ehaced TIC. Use these values to select regios of iterest for your TIC. You may wat to :- Look at a certai peak Igore solvet peaks Igore grassy baselies Not cout saturated peaks Note for Max Itesity, ios produce a electroic sigal betwee 0 ad 10 volts. The default value show here represets 10 volts. Pressig the Default butto loads default values for all parameters ad switches off all the advaced istrumet data thresholdig features. Press the OK butto to accept ay chages. The settigs will be dowloaded whe you press the OK butto i the Istrumet Data Thresholdig page. These settigs will ot be dowloaded if Cacel is pressed i the Istrumet Data Thresholdig page. Page 16

21 Chagig Lab ad User Iformatio MassLyx ca save maagemet iformatio with a data file. The iformatio saved is Laboratory Name, Istrumet Idetificatio ad User Name. This iformatio is etered ito the Lab ad User Ifo dialog box. This iformatio will be stored with ay data that is acquired ad ca be displayed as part of the header iformatio of a chromatogram or spectrum whe they are displayed o the scree or prited. Figure 2.10 Chage Maagemet Iformatio Dialog To chage Lab ad User Iformatio 1. Choose Chage Lab or User Ifo from the Acquisitio Cotrol Pael Cofigure meu. 2. Make required chages to the iformatio. 3. Press the OK butto. Other Meu Items O the Acquisitio Cotrol Pael Istrumet meu there are three optios used for diagostics. Trasputer Versio Choose this optio to display the versio umber of the Trasputer code ad the Mass Rage of the Mass Spectrometer. Verify Serial Comms Choose this optio to check the commuicatios betwee a Autosampler or GC ad the Mass Spectrometer. Trace Choose this optio to geerate a log file of all evets betwee MassLyx ad the trasputers. Do ot use this optio uless advised to by Micromass as it geerates very large data files. A tick mark appears ext to the optio if it is selected. Page 17

22 Page 18

23 Notes Page 19

24 Notes Page 20

25 Istrumet Tuig Itroductio Chapter 3 The Tue Page Before acquirig data it may be ecessary to check the tuig coditios of the istrumet, ad if ecessary modify oe or more of the istrumet tuig parameters. The istrumet ca be tued either maually or automatically from the istrumet Tue page. Figure 3.1 Tue Page showig tue peak iformatio The Tue page is a cofigurable paeled display as show i Figure 3.1. The left side of the widow holds the tuig parameters for a selected regio of the mass spectrometer. The Mass Spectrometer regio ca be chaged usig either the Widow meu, or by pressig oe of the buttos at the bottom left of the widow. The pael i the top right of the widow ca display either the tue peak iformatio (Figure 3.1 ) or istrumet pressure iformatio (Figure 3.2 ). The display ca be switched betwee tue peak iformatio ad istrumet pressure iformatio usig the Widow meu or the ad toolbar buttos. A Toolbar is displayed at the top of the tue widow, which allows you to perform some commo operatios with a sigle click of the appropriate Toolbar butto. Note the Toolbar does ot appear for some older istrumets like the , Trio-2000, Bio-Q ad Quattro I. Page 21

26 At the bottom right of the widow is the tue peak display. You ca display up to four masses to tue o. The umber of tue peaks displayed is cotrolled by the four check boxes at the bottom right-had side of the widow. Ay oe of the tue peaks ca be zoomed so that it occupies the etire tue peak area. Whe a tue peak has bee zoomed the cotrols for the mass ad spa for that peak are displayed at the top of the display widow. This eables you to display the pressure iformatio while havig cotrol over the peak display. Figure 3.2 Tue Page showig istrumet pressure iformatio To display the Tue Page 1. Select the Acquisitio Cotrol Pael. 2. Double click o the mass spectrometer sectio of the istrumet picture o the Acquisitio Cotrol Pael widow. -or- Choose Tue Mass Spectrometer from the Acquisitio Cotrol Pael Istrumet meu. 3. From the widow meu, select either Peak Cotrols to show peak iformatio or Pressures to show istrumet pressure iformatio. Page 22

27 The Tue Page Toolbar The Toolbar is displayed at the top of the tue widow ad allows you to perform some commo operatios with a sigle click of the appropriate Toolbar butto. Press to ope a existig tue file. Press to save curret tue parameters to disk. Press to prit curret widow i portrait format. Press to display tue peak iformatio. Press to display istrumet pressure iformatio. Press to edit scope settigs. Press to reset the zero level of the istrumet ad reiitialise the system. Press to ru AutoTue. Press oce to switch referece gas o. Press agai to switch referece gas off. Press oce to switch CI gas o. Press agai to switch CI gas off. Press oce to switch API gas o. Press agai to switch API gas off. Press oce to switch collisio gas o. Press agai to switch collisio gas off. Press to get help. Page 23

28 Chagig Tue Parameter Settigs Most of the tuig parameter cotrols are combiatio cotrols featurig a slider bar, two push buttos ad a edit widow. The tuig parameters ca be modified i ay of the followig ways. By draggig the slider bar usig the mouse. By clickig the "<" or ">" buttos. By typig a ew value directly ito the edit widow. If you chage a tue parameter usig the slider bar or push buttos, the value show i the edit widow will update as appropriate. Other tue parameter cotrols oly have a edit widow ad ca oly be chaged by direct typig. The speed with which the system will respod to chages i parameter settigs is cotrolled by the speed with which the peak display refreshes. To get the sliders to be most resposive you should set the scope sca speed ad iter sca time to be as short as possible (for more iformatio, see Scope Parameters, o page 31). Pritig Tue Iformatio A report of the tuig parameters ca be set to the priter by pressig the toolbar butto or by choosig Prit from the tue page File meu. This report cotais a copy of the tue peak iformatio displayed o the scree, alog with a record of each parameter settig. This report is ot cofigurable by the user. Experimetal Record Tuig parameters are stored with every data file as part of the experimetal record. The tuig parameters for a particular data file ca be viewed or prited from the Data Browser, see the MassLyx User Guide, Selectig ad Viewig Data, for more iformatio. Page 24

29 Savig ad Restorig Parameter Settigs Whole sets of istrumet tuig parameters ca be saved to disk as a amed file ad the recalled at a future date. Note: A tue parameter file cotais the latest settigs for the source cotrols for all supported ioizatio modes ot just the ioizatio mode you are curretly usig. Tue parameter files also cotai settigs for the aalyser, ilet set poits ad peak display. Save o Exit Optio Whe the Save o Exit optio is selected the curret tuig parameters are saved to disk whe you exit from the tue page ad will be reloaded whe you eter the tue page agai. Whe Save o Exit is ot selected the curret tuig parameters are ot saved to disk whe you exit from the tue page, whe you eter the tue page agai the last saved set of parameters will be reloaded from disk. To eable the Save o Exit optio, choose Save o Exit from the tue page File meu. A tick mark will appear ext to this optio to show that it is eabled. To disable the Save o Exit optio, choose Save o Exit from the tue page File meu agai. To save a set of parameters 1. To save the curret tue parameters with the existig file ame press the butto or choose Save from the Tue Page File meu. To save the curret tue parameters with a ew file ame choose Save As from the Tue Page File meu. 2. Eter a ew file ame uder which you wat the parameters to be saved, or alteratively select a existig file from the list displayed. 3. Press the Save butto. If the selected file already exists o disk you will be asked to cofirm that you wat to overwrite the existig iformatio. Press the Yes butto to cotiue or the No butto to eter a differet file ame. Figure 3.3 Save Tue Parameters Dialog Page 25

30 To restore a saved set of parameters 1. Press the toolbar butto or choose Ope from the Tue File meu. 2. Select the tuig parameter set that you would like to use, either by typig its ame or by selectig it from the displayed list. 3. Press the Ope butto. Figure 3.4 Ope Tue File Dialog Page 26

31 Modifyig the Peak Display The tue peak display is modified usig either the tue peak cotrols, or by usig the mouse directly o the tue peak display. Figure 3.5 shows the tue peak cotrols ad the tue peak display. Figure 3.5 Tue peak cotrols ad display To select peaks 1. Choose the umber of peaks that you wat to display by checkig the appropriate cotrols at the bottom of the widow. For example, if you wat to display oly the first ad secod tue peaks the check cotrols 1 ad 2 o, 3 ad 4 off. 2. Press the toolbar butto or choose Peak Display from the Tue Page Widow meu ad the for each active peak select the sca type (Quattro oly) that you wish to use, the mass that you wish to tue o, the spa ad the gai. To zoom or uzoom a peak Double click o the peak. This will make the selected peak fill the whole tue peak display ad remove the peak selectio cotrols from the widow. A secod double click will uzoom the peak. Page 27

32 To chage the tue mass, spa or gai usig the mouse 1. Zoom the relevat peak - this will cause left ad right arrow cotrols for the mass ad spa to appear at the top corers of the peak widow. 2. Click o the arrow to the left of the mass cotrol to decrease the mass by oe icremet, click o the arrow to the right to icrease the mass. The size of the icremet is set usig the scope cotrols dialog (see later). 3. To double the gai applied to a peak double click o the lie above the peak which shows the gai. A double click below the peak will half the gai. AutoTue MassLyx ca automatically tue the mass spectrometer i EI, APcI ad electrospray ioizatio modes o supported istrumets. The istrumets supported for each ioizatio mode are as follows: Autotue Ioizatio Mode EI APcI ad Electrospray Istrumets Supported Platform II, Quattro II, Quattro I ad Trio-2000 Platform II, Quattro II, Platform LC ad Quattro LC AutoTue ramps the settigs for the tuig parameters util they are optimised to give the best itesity, resolutio ad peak shape. To ru AutoTue 1. Press the toolbar butto or choose AutoTue from the Tue page Other meu. Note, the istrumet must be i operate before you ca ru AutoTue. Figure 3.6 AutoTue dialog Page 28

33 2. Choose Setup to set the AutoTue setup parameters. Figure 3.7 EI AutoTue Setup dialog The default setup parameters for EI AutoTue are suitable for tuig with Heptacosa, there is o eed to alter the Tue Mass or Peak Width parameters if you are usig Heptacosa. The Level of AutoTue performed ca be selected as Maiteace or Full. A full AutoTue starts from a default set of tuig parameters. A maiteace AutoTue starts from the curret tuig parameters set i the tue page ad ca be quicker tha a full AutoTue. A maiteace AutoTue ca oly be performed if the istrumet is reasoably well tued already, if the curret tuig is too poor AutoTue will give a error ad ask you to perform a full AutoTue. The Tue Masses parameters set a Low ad High Mass which will be used to tue o. The Peak Width at Half Height parameters set the peak width at half height for the Low ad High Masses beig moitored. Whe you are satisfied with the AutoTue setup parameters press OK to exit. 3. Choose Start to start AutoTue. The AutoTue status bar is updated to show the progress of AutoTue. AutoTue has 7 stages: 1) Checkig istrumet coditios Autotue checks that tue page readbacks are withi acceptable tolerace. 2) Attemptig to detect iitial beam AutoTue looks for a iitial beam by icreasig the value of the multiplier. Page 29

34 3) Rough tuig focus leses at low mass Values of leses are ramped to see where the low mass peak itesity maximises. 4) Tuig io eergy ad io repeller Modifies values for io eergy ramp, io eergy ad repeller. 5) Tuig for good peak shape Checks peak width at half height ad height of valley betwee mai ad isotope peaks. 6) Fie tuig focus leses at high mass Values of leses are ramped over a arrower rage usig the values determied i stage 4 to see where the high mass peak itesity maximises. 7) Tuig multiplier for good sesitivity Value of multiplier ramped util the low mass peak itesity is 80% of full scale. 4. Whe AutoTue has fiished it displays a status dialog to say that AutoTue has bee successfully completed. Press OK to exit back to the tue page. The tuig parameters determied by AutoTue will be saved to the curret tue parameter file. Page 30

35 Scope Parameters Various parameters ca be set to cotrol the peak display ad, if fitted, a exteral oscilloscope. You ca cotrol the speed at which the scope is refreshed ad the scope respose. You ca also ask for a grid to be applied to the tue peak display ad for the data to be draw either as a filled or ufilled trace. Figure 3.8 Scope parameters dialog To chage the scope setup 1. Press the toolbar butto or choose Scope Parameters from the Tue Other meu. 2. Make ay required chages to the settigs. 3. Press the OK butto. Respose A variable low pass filter that effects the level of high frequecy iformatio that will be displayed o a exteral oscilloscope, if oe is fitted to the mass spectrometer. A high value will make the peak display appear smoother tha a low value. Sca Time ad Iter Sca Time Cotrol the speed with which the tue peak display is updated. The tuig system will behave more resposively if the sca time ad iter sca time are short. Mass Icremet ad Spa Icremet The mass ad spa settigs for a zoomed peak ca be altered usig the mouse by clickig o the appropriate up or dow butto (see: Modifyig Peak Display). The mass icremet ad spa icremet cotrols effect how much the settig will chage for each mouse click. Grid Display a horizotal grid o the peak display. Page 31

36 Solid Trace Check this cotrol to draw the displayed peak as a filled trace. Leave the cotrol uchecked to draw the displayed peak as a lie graph. Peak Itesity Whe this parameter is set to Percetage the percetage itesity of the base peak i the active tue segmet widow is displayed. Whe this parameter is set to Abs Itesity the absolute itesity of the base peak i the active tue segmet widow is displayed. Chagig Ilet Heaters Figure 3.9 shows the dialog box that is used to set ilet iterface temperatures ad the DCI curret setpoit. The Heaters dialog is a "modeless" dialog, which meas that you do ot have to close it to cotiue usig the rest of the system. Figure 3.9 Ilet Heaters dialog for iterface temperature ad DCI curret setpoit To chage a ilet parameter 1. Select Chage Heaters from the Tue Heaters meu. 2. Select the required parameter ad eter the desired set poit i the edit box labeled Set To. 3. To sed the curret values to the istrumet press the Update butto. The ew set poit will ow be set to the acquisitio system. You ca observe the effect of the chage by watchig the read back value which should chage as the selected heater heats up or cools dow to the required value. Readbacks are displayed i the box labeled Curretly. 4. Press the OK butto to sed curret values to the istrumet ad exit from the Heaters dialog. Page 32

37 Gas Cotrols The Gas meu o the Tue page lets you tur gasses o or off. You ca also customize the meu so that the correct ames of the gasses that you actually use appear i the meu bar. To tur a gas o or off Press the relevat toolbar butto or choose the required gas from the Tue Gas meu. If the gas was previously tured off it will ow be tured o. A tick mark will appear ext to a gas if it is tured o. To chage the ame of a gas 1. Select Parameters from the Tue Gas meu. 2. Select the required gas ad eter its ame. If you wish to be able to select the gas from the Tue Gas meu usig a key letter you should precede this letter of the ame by the character "&". For example, i Figure 3.10 the CI Gas will appear o the Tue Gas meu as CI Gas ad ca be selected via the keyboard by pressig "I". Note the key letter does ot ecessarily have to be the first letter of the word. Figure 3.10 Tue gas parameters dialog Page 33

38 Ramp Cotrols For istrumets, which have the relevat hardware, istalled it is possible to set coe voltage ad collisio eergy ramps o the tue page. To set up a coe voltage ramp 1. Choose Coe Ramp Gradiet from the tue page Ramps meu. Figure 3.11 Coe Ramp dialog 2. Two values of coe voltage are defied at two particular masses usig the Start ad Ed Mass cotrols ad the Coe Start Volts ad Coe Ed Volts cotrols. These values defie a gradiet for the coe voltage which is the extrapolated to cover the full mass rage. Make ay chages required ad press OK to exit. Pressig the Update butto i this dialog will update the values used for the coe voltage ramp whe the ramp has bee eabled. 3. To iitiate the coe voltage ramp press the butto or choose Use Coe Ramp from the tue page Ramps meu. To set up a collisio eergy ramp 1. Choose Collisio Eergy Ramp Gradiet from the tue page Ramps meu. Figure 3.12 Collisio Eergy Ramp dialog 2. Two values of collisio eergy are defied at two particular masses usig the Start ad Ed Mass cotrols ad the Collisio Eergy Start Volts ad Collisio Eergy Ed Volts cotrols. These values defie a gradiet for the collisio eergy voltage that is the extrapolated to cover the full mass rage. Make ay chages required ad press OK to exit. Pressig the Update butto i this dialog will update the values used for the collisio eergy ramp whe the ramp has bee eabled. 3. To iitiate the collisio eergy voltage ramp press the butto or choose Use Collisio Eergy Ramp from the tue page Ramps meu. Page 34

39 Vacuum The mass spectrometer s vacuum system ca be cotrolled from the Tue page. Esure however that this is doe i accordace with the iformatio i your Istrumet Users Guide. To pump dow the vacuum system Choose Pump from the Tue Other meu. The meu ame will chage from Pump to Vet ad the system will commece its pump dow sequece. You ca watch the vacuum progress by choosig Pressures from the Tue Widow meu. To vet the vacuum system Choose Vet from the Tue Other meu. You will be asked to cofirm that you wat to vet the system, i case Vet was selected i error. The system will the start its automatic vetig sequece. Resettig the Zero Level The zero level (or Baselie) ca be repositioed by Pressig the toolbar butto or choosig Reiitialize from the Tue Other meu. This commad will cause the istrumet cotrol system to measure the positio of the oise sigal so that ay baselie offset caused by the electroics or istrumetatio ca be compesated for. It is advisable to reset the zero level wheever the value of oe of the multiplier voltages is chaged. Page 35

40 Cotrollig Readbacks MassLyx allows you to choose how system readbacks will be displayed o the Tue page. You ca ask for readbacks to be displayed cotiuously, ever or oly whe they differ from their associated set poits by more tha 10%. Note that a umber of the readbacks are for diagostic purposes oly ad do ot eed to be very precise. Their fuctio is to idicate if the voltage is preset or ot. The acceptable variatio betwee the set value ad the readback value will vary depedig o the particular tue parameter. If you are cocered about the values you should cotact your local service office for advice. To chage read back style 1. Choose Readbacks from the Tue Other meu. 2. Select the readback style required. Selectig Always will always display readbacks. Selectig Out of Rage will display readbacks whe they differ from their associated set poits by more tha 10%. If both cotrols are uchecked the o readbacks will be displayed. 3. Press the OK butto. Figure 3.13 Tue Readbacks dialog Page 36

41 Startig a Acquisitio from the Tue Page The easiest way to acquire data for a sample is to acquire it directly from the tue page. You caot use ilet programs from the tue page, acquire aalog data or acquire multiple sample sequeces, but you ca start ad stop acquisitios ad cotrol most of the scaig parameters. The Start Acquisitio dialog as show i Figure 3.14 ca be folded to hide most of the parameters from the user ad ca be cofigured to automatically set itself to acquire data usig the mass rage ad fuctio type that is beig used for tuig the istrumet. These features provide a WYSIWYG (What you see is what you get), method of acquirig data as the spectrum that is writte to disk will be the same as the spectrum that is show i the peak display. Figure 3.14 Start Acquisitio dialog To start a acquisitio 1. Choose Acquire from the Tue Widow meu or click o the Acquire butto at the bottom of the tue widow. 2. Make ay required chages to the settigs. 3. Press the Start Acquisitio butto. Fileame The ame of the data file to which acquired data will be writte. The fileame ca be up to 128 characters. If the file already exists o disk, you will be asked if you wat to overwrite it. Page 37

42 Text A text area that is used to eter the sample descriptio. The descriptio ca be displayed o ay output of the acquired data ad has a maximum legth of 80 characters. Directory The Directory field displays the directory ito which data will be acquired. If you wish to chage the directory ito which data will be acquired, you ca cacel the acquisitio ad create a ew project by choosig Project Wizard, or ope a existig oe by choosig Ope Project, from the MassLyx Top Level File meu. Origi Pressig the Origi butto allows you to eter additioal iformatio about the sample to be aalysed. Additioal iformatio ca be etered ito the followig fields Submitter, Job, Task ad Coditios. Fuctio The sca fuctio that will be used to acquire the data. It ca be ay of the followig MS, Paret, Daughter, Neutral Loss, Neutral Gai, Q1F, MS2. More iformatio is give o sca fuctios later o i this chapter. Data Format The type of data that will be collected ad stored o disk. It ca be ay of the followig Cetroid, Cotiuum or MCA. More iformatio is give o data formats later o i this chapter. Save o Exit If this cotrol is checked, ay chages that you make will be saved whe you exit the dialog. Use Scope Settigs If this cotrol is checked, the mass iformatio ad fuctio type will be take directly from the curretly zoomed peak o the tue page Set Mass Specifies the mass ( i.e. Daughter Mass, Paret Mass etc. ) that will be used for the particular fuctio type. If the fuctio that you have selected does ot require a set mass, this cotrol will be disabled. Start Mass Specifies the mass at which the sca will start. The Start Mass must be lower tha the Ed Mass. Ed Mass Specifies the mass at which the sca will stop. Scope Gai The amout of gai that will be applied to data displayed o a exteral oscilloscope uit, if oe is fitted. Sca Time Specifies the duratio of each sca i secods. Iter Sca Time Specifies the time i secods betwee a sca fiishig ad the ext oe startig. Durig this period o data is stored. Ru Duratio Legth of the acquisitio, measured i miutes. Page 38

43 Q-Tof Tuig Q-Tof machies have extra parameters to set which are accessed from the Tue Page Other meu. To display the relevat dialog select the optio from the meu. TDC Settigs Figure 3.15 TDC Settigs dialog Start Eter the start threshold for the TDC sigal. Stop Eter the stop threshold for the TDC sigal. Threshold Eter a itesity threshold below which TOF data will ot be stored. Lteff This is used to adjust the mass scale. Page 39

44 RF Settigs Figure 3.16 RF Settigs dialog Eter a Gai ad Offset for the Trasport RF ad Collisio RF. Update Pressig this butto seds the values to the istrumet without closig dow the dialog. MCP Coditioig Figure 3.17 MCP Coditioig dialog Eter the Start ad Stop voltages, the Duratio ad Step values. The MCP voltage will be stepped betwee the specified voltages for the specified time. Whe the Start butto is pressed the MCP Coditioig Status dialog is displayed. Press the Stop butto to stop MCP Coditioig. Page 40

45 Figure 3.18 MCP Coditioig Status dialog TOF MS Profile Figure 3.19 TOF MS Profile dialog This dialog allows the user to bias differet parts of the mass rage durig TOF MS acquisitio. The Quadrupole will park at the specified Mass for the specified percetage of the sca time (Dwell Time), ad the ramp to the ext mass for the specified percetage of the sca time (Ramp Time). Update Pressig this butto seds the values to the istrumet without closig dow the dialog. Page 41

46 Page 42

47 Notes Page 43

48 Notes Page 44

49 Mass Calibratio Mass Calibratio Chapter 4 MassLyx provides a fully automated facility for calibratig the mass scale of a istrumet. To see the mai calibratio dialog choose Calibrate.. from the acquisitio cotrol pael Istrumet meu. How a calibratio is formed Calibratio Types A mass spectrum of a referece compoud ( a calibratio file ) is acquired ad matched agaist a table of the expected masses of the peaks i the referece compoud which are stored as a referece file. Each peak i the referece file is matched to a correspodig peak i the calibratio file. The mass differeces betwee the referece peaks ad calibratio peaks are the calibratio poits. A calibratio curve is fitted through the calibratio poits. The vertical distace of each calibratio poit from the curve is calculated. This distace represets the remaiig ( or residual ) mass differece after calibratio. The stadard deviatio of the residuals is also calculated. This umber is the best sigle idicatio of the accuracy of the calibratio. A sigle-quadrupole istrumet requires up to three calibratio curves; A static calibratio is used to accurately park the quadrupole mass aalyser o a specific mass of iterest (i Tuig ad SIR for example). A scaig calibratio eables peaks acquired i a scaig acquisitio to be mass measured accurately A sca speed compesatio calibratio compesates for lag time i the system whe the istrumet is scaed rapidly. A separate mass spectrum of the referece compoud is acquired for each selected calibratio type. A triple-quadrupole istrumet requires these three calibratios for both MS1 ad MS2, for a maximum of six calibratio curves. The table below show which types of calibratio are ecessary for particular types of experimet. Page 45

50 Experimet Calibratio Required MS1 MS2 MS All Not Applicable SIR Static Not Applicable MSMS All All MRM Static Static Overview of the calibratio process 1. Check the tuig of the istrumet. The mass spectrometer should be i operatioal mode. Check that the peak shape ad itesities are correct. 2. Set the calibratio parameters. Select the appropriate referece file for the calibratio referece sample that you are goig to use. If you are calibratig electrospray, FAB or thermospray, the you will typically be usig cotiuum or MCA data types, ad you should set the Mass Measure parameters (see page 52). These parameters cotrol peak detectio i cotiuum ad MCA data. There is o eed to set these parameters if you are usig cetroided acquisitio. Set the Peak Match ad Curve Fit parameters i the Calibratio Parameters dialog (see page 50). These parameters cotrol the locatio of referece peaks i a calibratio spectrum, ad the drawig of a calibratio curve to correct the resultig mass differeces. 3. Start a automatic calibratio From the Mai Calibratio Dialog, press the Start... butto. This will brig up the automatic calibratio dialog. Select the types of calibratio you wish to perform. Check the Acquire ad Calibrate checkbox to perform automatic calibratio. Set the stadard deviatio threshold. Optioally, also check the Acquire ad Verify checkbox, ad set the stadard deviatio threshold. Check the Prit Report checkbox if you wat a prited calibratio report. Set the data acquisitio parameters. Press the OK butto. Page 46

51 4. Check the calibratio report Examie the calibratio report, ad display the calibratio graphs if ecessary, to satisfy yourself that the calibratio is satisfactory. The Calibratio Dialog The calibratio dialog displays status iformatio, icludig calibrated mass rage ad sca speeds, ad the time ad date of the last calibratio. To display the dialog select Calibrate from the Acquisitio Cotrol Pael Istrumet meu. Figure 4.1 Mai Calibratio dialog The dialog may be folded to hide the status iformatio by pressig the <<Fold butto. Whe i the folded state, the status iformatio may be displayed by pressig the Status>> butto. Figure 4.2 Mai Calibratio dialog (ufolded triple quad versio) Page 47

52 Parameters which cotrol Calibratio Automatic calibratio parameters To access this dialog, press the Start butto o the Calibrate dialog. Figure 4.3 Automatic Calibratio dialog You may choose Static Calibratio, Scaig Calibratio, or Sca Speed Compesatio calibratios by checkig the appropriate check boxes. O a triple quadrupole istrumet you must also specify MS1 ad/or MS2. Optioally, you may choose to Verify the calibratio. Selectig this optio causes a calibrated mass spectrum of the referece compoud to be acquired. Fially you ca choose to have a report of the calibratio prited automatically whe the calibratio completes. If you choose ot to prit the report at this stage you ca always prit it from the calibratio curve display later. The report cotais pictures of the calibratio curves produced, alog with calibratio statistics such as stadard deviatio. Note, for Q-Tof machies Types available are Quad ad TOF. Quad performs a stadard MS1 calibratio, TOF performs a Tof MS data calibratio. Select oly oe of these optios. Page 48

53 Data Acquisitio Parameters To chage data acquisitio parameters press the Acquisitio Parameters butto o the Automatic Calibratio dialog. Figure 4.4 Calibratio Acquisitio Setup dialog The Sca From ad Sca To parameters specify the sca rage for each calibratio type. The Ru Duratio parameter specifies how log each calibratio data file will be acquired for. The Data Type parameter specifies whether data will be acquired i cetroid, cotiuum or MCA mode. It is recommeded that the calibratio uses the same data type as the sample data you will be acquirig. To acquire data for static calibratio, the portio of the mass scale immediately aroud each referece peak is scaed. The Static Spa parameter specifies the distace to be scaed either side of the referece peak. The Static Dwell parameter specifies the time take to sca this rage. To acquire data for scaig calibratio, the mass scale is scaed over the selected rage, i a time specified by the Slow Sca Time parameter. To acquire data for the Sca Speed compesatio calibratio, the mass scale is also scaed over the selected rage, i a time specified by the Fast Sca Time parameter. The Iter Sca Delay specifies the time betwee oe sca edig ad the ext sca startig. Pressig the Default butto will reset all parameters i the dialog box to their default values. Page 49

54 To chage Calibratio Parameters 1. Select Calibrate from the Acquisitio Cotrol Pael, this displays the Calibrate dialog. 2. Select Calibratio Parameters from the Edit meu of the Calibrate dialog. Figure 4.5 Calibratio Parameters Dialog Perform auto peak matchig Check the box to eable. Whe uchecked this allows disablig of the automatic peak matchig facility. Useful for TOF calibratio, where the automatic peak matchig algorithm will typically ot make the correct assigmets. The Peak Widow parameter specifies the maximum mass differece betwee the remaiig referece peaks ad the expected positio of the correspodig peaks i the acquired spectrum. Normal operatig rage for the Peak Widow parameter is 0.3 to 1.5 Da. The first referece peak to be matched is chose to be close to the ceter of the calibratio mass rage. The Iitial Error parameter specifies the maximum mass differece betwee this referece peak ad the correspodig peak i the acquired spectrum. Normal operatig rage for the Iitial Error parameter is 0.5 to 2.0 Da. Ay peak i the acquired spectrum with itesity less tha the specified Itesity Threshold will ot be used to form the calibratio curve. The threshold is specified as a percetage of the most itese peak i the acquired spectrum. Normal operatig rage for the Itesity Threshold parameter is 0 to 5%. Whe each peak i the referece spectrum has bee matched with a correspodig peak i the acquired spectrum, the mass differece acquired mass - referece mass is calculated for each pair of peaks. These mass differeces are plotted as poits o a graph; each data poit has the mass of the acquired peak as its x co-ordiate, ad the above mass differece as its y co-ordiate. A smooth curve is draw through the Page 50

55 poits. The Polyomial Order parameter cotrols the type of curve which is draw ad ca be set to ay value betwee 1 ad 5. If Polyomial Order = 1, a straight lie is draw through the poits. If Polyomial Order = 2, a quadratic curve is draw through the poits. If Polyomial Order = 3, a cubic curve is used. For a typical EI calibratio where the mass rage calibrated starts below 100 amu ad exteds up to 650 amu the recommeded settig for the Polyomial Order parameter is 4. For a typical electrospray calibratio where the mass rage calibrated is from 600 amu to greater tha 1000 amu the recommeded settig for the Polyomial Order parameter is 2. Itesity Weightig If this box is checked, the curve fit is weighted toward the poits represetig the more itese acquired peaks. The weight of each poit is proportioal to the square root of the itesity of the acquired peak. Calibrate display Eables calibratio of the peaks i the top (raw file) graph. This feature allows selectio of oe peak at a time, with the display beig recalibrated after the selectio of each peak, brigig the other masses i the spectrum ito lie. Agai, useful for TOF calibratio. Automatic Calibratio Check parameters To access this dialog, select AutoCal Check Parameters from the Edit meu o the Calibrate dialog. Figure 4.6 Automatic Calibratio Check dialog The Missed Referece Peaks cotrol allows you to eter a umber of cosecutive peaks from the referece file that the system is allowed to miss before a warig is issued to the user. The Maximum Stadard Deviatio cotrol does the same thig if the residuals for a particular calibratio exceeds the umber etered. The Apply Spa Correctio optio causes a extra correctio to be applied to the mass scale, which is depedet o the mass rage beig scaed. This correctio esures that mass assigmet will be correct eve if the mass scale that you are workig with is differet to the oe that the istrumet was origially calibrated over. It is ot recommeded that this optio be used if the mass rage of iterest is less tha 1000amu ad icludes the rage amu. The Check Acquisitio Calibratio Rages optio causes a warig message to be displayed if a acquisitio is started which would be outside the rage of the curret calibratio. Page 51

56 The Mass Measure parameters If you are usig cotiuum or MCA acquisitio modes to acquire your calibratio data you will eed to tell the system how it should covert the acquired data ito cetroided data eeded by the calibratio process. To access this dialog, select Mass Measure Parameters from the Edit meu o the Calibrate dialog. QTOF Figure 4.7 Mass Measure dialog For a full descriptio of the Mass Measure parameters ad their effect, see the sectio headed "Mass Measure" i the Spectrum chapter of the MassLyx User Guide. For Q-Tof data this dialog will have a extra butto. Press this butto to display the QTOF Accurate Mass parameters dialog. For details see QTOF Accurate Mass below. There is also a extra field Use QTOF mass correctio. Check this box to use QTOF mass correctio. Page 52

57 QTOF Accurate Mass Figure 4.8 QTOF Accurate Mass dialog Resolutio Eter the resolutio of the Mass Spectrometer. Np multiplier Eter a value for the umber of pushes correctio factor. Mass Widow This parameter determies the width of the mass widow used to locate the lock mass data peak. The most itese peak i the rage Lock Mass Mass Widow to Lock Mass + Mass Widow is selected, ad mass correctio based o this peak is performed. Lock Mass This parameter specifies the referece lock mass. Page 53

58 The Calibratio Report If the calibratio proceeds successfully, o output will be displayed util the calibratio report is prited, showig calibratio curves for each selected calibratio type. If the stadard deviatio of the residuals for a particular calibratio exceeds the preset maximum, a set of calibratio graphs will be displayed to help idetify the problem. Typically, the peak matchig algorithm has foud the wrog peaks, or missed some peaks. You may eed to chage the peak match parameters or adjust the peak matchig maually. You ca chage the peak match parameters by choosig Peak match params from the calibratio graph Edit meu. If the peak matchig algorithm has foud the wrog peaks, icreasig the value of the Peak widow parameter will solve the problem. You should also try reducig the value of the Itesity threshold parameter (if it is ot already zero), before idetifyig the peaks i the acquired spectrum by had. Figure 4.9 Calibratio Graphs The top graph shows the calibratio spectrum; the peaks matched to referece peaks are highlighted i a differet color. Page 54

59 Alterig the displayed rage You ca alter the rage of the spectrum o display by clickig the left mouse butto o a graph at oe ed of the rage of iterest, holdig the butto dow ad draggig the mouse to the other ed of the rage of iterest. A "rubber bad" idicates the selected rage. Pressig the Default butto restores the default display rage. Maually matchig peaks You ca match a calibratio peak to the closest referece peak by positioig the mouse poiter over the calibratio peak ad clickig the right mouse butto. If the closest referece peak is already matched to aother calibratio peak, the previous match will be removed. You ca also use the right mouse butto to udo a peak match, by positioig the mouse poiter over the matched calibratio peak ad clickig the right mouse butto. Other Calibratio Facilities To delete the istrumet calibratio 1. Choose Delete all calibratio from the Calibrate Process meu. This will brig up a dialog requestig cofirmatio. 2. Press OK to delete all calibratio, or Cacel to abort the operatio. Displayig a calibratio graph You ca display the calibratio graphs for a particular calibratio type as follows: 1. Choose Calibratio from file from the Calibrate Process meu. 2. Select Static or Dyamic sca, or Lag time calibratio for MS1 or MS2 (o a triple quad istrumet). 3. Press the OK butto. Page 55

60 Makig a calibratio from a data file Figure 4.10 MS1 Scaig Calibratio dialog 1. Choose Calibratio from file from the Calibrate Process meu. 2. Select Static, Scaig or Sca Speed Compesatio calibratio for MS1 or MS2 (o a triple quad istrumet). 3. Press the Browse butto, ad the select a file from the list displayed. 4. Optioally, eter a rage of scas to combie From ad To. 5. Press OK. Page 56

61 Editig a Referece File Figure 4.11 shows the cotets of the file Heptacos.ref which is the stadard referece for EI calibratio. Calibratio referece files cosist of two colums of umbers separated by ay umber of spaces or TAB characters. The first colum cotais the referece peak masses ad the secod colum cotais the referece peak itesities. The referece peak itesities are ot at preset used by the calibratio software ad so ca be set to a omial value of 100. However, you may wish to eter realistic values here to improve the appearace of the referece spectra Figure 4.11 Heptacos.ref file Calibratio referece files ca be created or edited usig ay Widows text editor. To read the curretly selected referece file ito the Notepad text editor, choose Referece File from the Calibratio Edit meu. After editig, the referece file ca either be saved uder the curret ame by selectig Save from the Notepad File meu or saved as a ew referece file by selectig Save as from the Notepad File meu ad givig the file a ew ame. Textual iformatio or commets ca be stored i the referece file. Lies, which are textual iformatio or commets, must start with the ; character. Savig ad Restorig Calibratios The complete istrumet calibratio ca be saved to disk as a amed file ad the recalled at a future date. Static, dyamic ad lag time calibratios are all saved together uder a commo ame. You may fid it useful to calibrate the istrumet for each of the differet types of experimet that you do ad save these calibratios to disk. This meas that whe you switch betwee experimets you ca restore a suitable calibratio from disk rather tha havig to recalibrate from scratch. Page 57

62 To save a amed calibratio 1. Choose Save Calibratio As from the Calibratio File meu. 2. Eter a ew file ame uder which you wat the parameters to be saved. 3. Press the Save butto. If the selected file already exists o disk you will be asked to cofirm that you wat to overwrite the existig iformatio. Press the OK butto to cotiue or the Cacel butto to eter a differet file ame. Figure 4.12 Save Calibratio Dialog To restore a saved calibratio 1. Choose Load Calibratio from the Tue File meu. 2. Select the calibratio file required, either by typig its ame or by selectig it from the displayed list. 3. Press the Ope butto. Figure 4.13 Load Calibratio Dialog Page 58

63 Notes Page 59

64 Notes Page 60

65 The Fuctio List Editor Itroductio Chapter 5 The Fuctio List Editor is used to set up the fuctio(s) that the mass spectrometer will use to sca the istrumet durig a acquisitio. A fuctio list ca be a mixture of differet scaig techiques that ca be arraged to ru either sequetially or cocurretly durig a acquisitio. Typical uses for mixed fuctio acquisitios are to acquire differet SIR groups over differet retetio widows ad the ability to switch MS/MS sca modes durig a acquisitio. A fuctio list is produced, saved o disk ad the refereced by ame whe you start a acquisitio. Figure 5.1 shows a simple fuctio list that cotais oly oe fuctio: a cetroided mode full sca, betwee 50 ad 550 amu usig EI ioizatio. Immediately above the fuctio bar display is a time scale that shows whe the fuctio will be active from ad for how log it will ru. I this case the fuctio starts after 5 miutes ad the rus for 35 miutes, termiatig after a total elapsed time of 40 miutes. To access this dialog, press the toolbar butto o the MassLyx Scree or select Mass Spectrometer from the Methods meu o the Acquisitio Cotrol Pael. Figure 5.1 Fuctio list showig a sigle fuctio The curretly selected fuctio is eclosed i a rectagular frame. If the display shows more tha oe fuctio you ca select a ew fuctio either by clickig o it with the mouse, or by movig to it usig the Up or Dow Arrow keys o the keyboard. Page 61

66 Figure 5.2 shows a more complicated fuctio list, which has four SIR fuctios, ruig sequetially for 10 miutes each. Figure 5.2 Fuctio list showig four SIR fuctios Addig a New Fuctio A ew fuctio ca be added either by clickig o oe of the fuctio buttos at the top of the editor, or by selectig Add from the Fuctio meu ad choosig the appropriate fuctio. The editor for the fuctio type selected will be displayed showig default values. Make ay chages required to the parameters ad press the OK butto to add the ew fuctio. The fuctio editors for each sca type will be discussed i detail later o i this chapter. Modifyig a Existig Fuctio A existig fuctio ca be modified i oe of two ways. You ca either choose Edit Fuctio from the Fuctio meu or you ca double click o the fuctio i the fuctio list. This will brig up the appropriate editor for the fuctio type ad allow you to chage the fuctio iformatio. Whe you have fiished editig the fuctio, the fuctio list display will be updated to show ay chages. Removig a Fuctio You ca remove a fuctio by highlightig it ad the choosig Delete Fuctio from the Fuctio meu. Alteratively you ca press the Delete key o the keyboard. You will be asked to cofirm that you really wat to delete the fuctio, press the Yes butto. Page 62

67 Chagig a Fuctio s Start ad Ed Times You ca chage the start ad ed times of a fuctio either by goig ito its editor as described above, or by pickig the times off a trace displayed i Chromatogram. To set a fuctio time from Chromatogram 1. I the fuctio list editor, select the fuctio that you wat to set. 2. Click with the right mouse butto ad drag over the required retetio widow o ay chromatogram trace i the Chromatogram widow. The curret fuctio bar will ow update to show the ew start ad ed times. Settig a Solvet Delay Aalog Chaels A solvet delay ca be set for a fuctio list usig the Solvet Delay Time cotrol. No data is stored durig the solvet delay period, which meas that solvet peaks that would ormally be see elutig at this time o the TIC chromatogram of the acquired data will o loger be see. The filamet i the source is tured off durig the solvet delay to prevet it from beig damaged. For APcI fuctios the APcI probe temperature will be set to the value specified i the APcI Probe Temp cotrol for the period of the solvet delay. If a aalog chaels hardware optio is fitted to your istrumet, you ca acquire up to 4 chaels of aalog data which will be stored with the data acquired from the mass spectrometer. Aalog chaels will typically be used to collect data from exteral uits such as UV detectors or FID detectors. A readig is made from the exteral chael at the ed of each sca ad stored with the data for that sca. The resolutio of the chromatography for a Aalog chael is therefore depedet o the sca speed used to acquire the mass spectrometry data. To access this dialog, select Aalog Chael Desc from the Aalog meu o the Sca Fuctios dialog. Figure 5.3 Aalog Chaels Descriptio dialog Page 63

68 To store data for a aalog chael 1. Check the boxes required i the Aalogue Chaels part of the fuctio editor. 2. Choose Aalog Chael Desc... from the Fuctio List Aalog meu. 3. Eter a textual descriptio for each of the aalog chaels that you have chose to store. This descriptio will be used o the Aalog Chromatogram dialog as the Chael descriptio. (See the MassLyx User s Guide, Chromatogram chapter). 4. Press the OK butto. Savig ad Restorig a Fuctio List To save a fuctio list 1. Choose Save As from the Fuctio List File meu, to brig up the Save File As dialog. 2. Eter a ew File Name uder which you wat the fuctio list to be saved, or alteratively select a already existig file from the list displayed. 3. Press the Save butto. If the selected file already exists o disk you will be asked to cofirm that you wat to overwrite the existig iformatio. Press the Save butto to cotiue, or Cacel ad select a differet ame. Figure 5.4 Save fuctio list dialog The curret fuctio list, i.e., the fuctio list that is o display i the fuctio list editor, is saved automatically to disk whe the fuctio list editor is closed. Therefore it is ot ecessary to save the fuctio list maually before closig the widow. The same fuctio list will be automatically loaded whe the fuctio list editor is ext accessed. Page 64

69 To restore a saved fuctio list 1. Choose Ope from the Fuctio List File meu to brig up a dialog similar to the oe show below. 2. Select the File Name of the fuctio list that you would like to use, either by typig its ame or by selectig it from the displayed list. 3. Press the Ope butto. Figure 5.5 Ope fuctio list dialog Page 65

70 Settig up a Full Sca Fuctio The full sca fuctio editor is used to set up cetroid, cotiuum ad MCA fuctios. It is activated by clickig o the MS1 fuctio butto. Figure 5.6 Full sca editor Start Mass Specifies the mass at which the sca will start. The Start Mass must be lower tha the Ed Mass. Ed Mass Specifies the mass at which the sca will stop. Both the start ad ed masses ca be selected directly from a spectrum displayed i the Spectrum widow. If you click, drag ad release o a spectrum usig the right had mouse butto, the selected mass rage will be etered as the start ad ed mass values. Ioizatio Mode Specifies the ioizatio mode ad polarity that will be used durig the acquisitio. Data Specifies the type of data to be collected ad stored o disk. Cetroid - Stores data as cetroided, itesity ad mass assiged peaks. Data is stored for every sca. Cotiuum - Data is ot cetroided ito peaks. Istead the sigal received by the iterface electroics is stored regularly to give a aalog itesity picture of the data beig acquired. Data is stored for every sca. Page 66

71 Due to the fact that data is beig acquired to disk at all times, eve whe there are o peaks beig acquired, cotiuum data acquisitio places some extra burde o the acquisitio system as compared to cetroided acquisitio. Data file sizes will ted to be sigificatly larger tha cetroided oes ad the absolute scaig speed (amu / sec) will be slower. It is possible however to set a threshold below which the data will ot be stored to disk, which ca greatly reduce these effects, depedig o the ature of the data beig acquired. The threshold ca be set so that data cosidered to be oise ca be discarded, thus improvig data acquisitio speed ad reducig data file sizes. For more iformatio about settig istrumet data thresholds see "Settig Istrumet Data Thresholds i Chapter 2. Multi Chael Aalysis (MCA) - MCA data ca be thought of as summed cotiuum, with oly oe itesity accumulated sca beig stored to disk for a give experimet. As each sca is acquired, its itesity data is added to the accumulated summed data of previous scas. A advatage of MCA is that radom oise will ot accumulate so rapidly as real data ad, therefore will effectively average out over a umber of scas. This will emphasise the real data ad improve sigal to oise. A further advatage of MCA is that because data is writte to disk oly at the ed of a experimet, scaig speeds ca be icreased ad sigificatly less storage space is required. The disadvatage of MCA is that as there is oly oe sca it caot be used for time resolved data. Repeats Is oly relevat for experimets havig more tha oe fuctio ad specifies the umber of repeats of this fuctio. Retetio Widow Start Time Specifies the retetio time i miutes at which this fuctio will become active; i.e., data acquired ad stored. Retetio Widow Ed Time Specifies the retetio time i miutes at which this fuctio will cease to be active; i.e., data acquired ad stored. Sca Time Specifies the duratio of each sca i secods. Iter-Sca Delay Specifies the time i secods betwee a sca fiishig ad the ext oe startig. Durig this period o data is stored. APcI Probe Temp Specifies the APcI probe temperature i degrees cetigrade. The APcI probe temperature cotrol is eabled whe the ioizatio mode is set to APcI. Use Tue Page Settigs Whe this cotrol is selected the APcI probe temperature set o the tue page at the start of the acquisitio is used. This cotrol is eabled whe the ioizatio mode is set to APcI. The APcI probe temperature value caot be altered by typig ew values ito tue page durig the acquisitio sice the ew values will ot be dowloaded durig the acquisitio. If you eed to alter the APcI probe temperature by typig ew values ito tue page durig the acquisitio you should acquire from the tue page. Coe Voltage Specifies the coe voltage value i volts. The Coe Voltage cotrol is eabled whe the ioizatio mode is set to ESP or to APcI. Page 67

72 Use Tue Page Settigs Whe this cotrol is selected the coe voltage set o the tue page at the start of the acquisitio is used. This cotrol is eabled whe the ioizatio mode is set to ESP or to APcI. The coe voltage value caot be altered by typig ew values ito tue page durig the acquisitio sice the ew values will ot be dowloaded durig the acquisitio. If you eed to alter the coe voltage by typig ew values ito tue page durig the acquisitio you should acquire from the tue page. Coe Voltage Ramp To apply a ramp to the coe voltage check the Use Coe Voltage Ramp cotrol ad press the CV Ramp... butto to load the Coe Ramp dialog. Figure 5.7 Coe Ramp dialog Two values of coe voltage are defied at two particular masses usig the Start ad Ed Mass cotrols ad the Coe Start Volts ad Coe Ed Volts cotrols. These values defie a gradiet for the coe voltage which is the extrapolated to cover the full mass rage of the fuctio. Page 68

73 Settig up a SIR Fuctio The SIR (Selected Io Recordig) techique is typically used i those situatios where oly a few specific masses are to be moitored durig a acquisitio. Sice most of the data acquisitio time is spet o these masses, the SIR techique is far more sesitive tha full scaig. The SIR editor is used to eter the masses that you would like to moitor ad their respective dwell time, spa ad iter-chael delay time. To set up a SIR fuctio Press the SIR butto o the Sca Fuctios dialog. Figure 5.8 SIR editor Most of the fields are the same as those described for the full sca editor (see "Settig up a full sca fuctio" o page 66). However the followig are differet: Mass Specifies the mass to be moitored; up to a maximum of 32. A mass ca be etered either by typig its value ito the Mass box ad pressig Eter or the Add butto, or by pullig them directly from a spectrum displayed i the Spectrum widow. To do this, display the required spectrum i the Spectrum widow ad the select the ios that you wish to moitor with a sigle click of the right had mouse butto. As you select a mass it will appear i the SIR masses table. Masses ca also be take directly from the sample list. Check the Use Sample List Masses box, ad eter sample list colum headig for the required masses (mass_a to mass_t) the press the Add butto. Note the colum headig (MASS_A, MASS_B etc) will appear i this dialog rather tha the actual masses listed i the sample list. Page 69

74 Dwell Specifies the legth of time i secods for which the highlighted mass will be moitored. Iter Chael Delay Specifies the legth of time i secods betwee fiishig moitorig the highlighted mass ad startig moitorig the ext mass i the fuctio. Spa Specifies a small mass widow applied cetrally about the highlighted mass. Durig acquisitio this rage will be scaed over the specified "Dwell" time. A spa of zero ca be set to simply sit o the specified mass. APcI Probe Temp Specifies the APcI probe temperature i degrees cetigrade. The APcI probe temperature cotrol is eabled whe the ioizatio mode is set to APcI. Use Tue Page Settigs Whe this cotrol is selected the APcI probe temperature set o the tue page at the start of the acquisitio is used. This cotrol is eabled whe the ioizatio mode is set to APcI. Add Type values ito the Mass ad Dwell ad the press this butto to add the ew Mass to the list. Chage Click with the left mouse butto o a mass o the list ad press this butto to chage the Mass or Dwell. Sort Press this butto to sort the list i order of ascedig Mass. Clear All Press this butto to delete the list of Masses. Delete Click with the left mouse butto o a mass o the list ad press this butto to delete a sigle Mass. Settig up a MS/MS Fuctio The MS/MS editor is used to acquire sca data usig the MS2 regio of a triple quadrupole mass spectrometer. If you have a sigle stage quadrupole istrumet you will ot be able to access this dialog box as it would be iappropriate to do so. The followig sca fuctios are available, ad for more iformatio you should refer to the Istrumet Users Guide. Daughter This is the most commoly used MS/MS mode ad is used to look at fragmetatios of a particular io. Quadrupole 1 is used to select the paret mass, ad is ot scaed. The mass for Quadrupole 1 is iserted ito the Paret mass edit widow. As a operatioal ote, the resolutio of Quadrupole 1 ca be lowered util the peak width at the base is two masses wide without the daughter spectrum cotaiig ay ios from the adjacet paret masses. The abudace sesitivity of the quadrupole is such that the umber of ios that are trasmitted at adjacet masses uder these coditios is very small ideed. Page 70

75 Figure 5.9 MS/MS editor for daughter sca The Start ad Ed masses refer to the mass rage to be scaed by Quadrupole 2. Note that it is possible to select the daughter mass to be greater tha the paret (precursor) mass. I this case ios which have gaied mass i the collisio cell, or are of higher mass to charge ratio will be detected. This occurs whe a multiply charged io fragmets ad loses a charge, but oly a fractio of the mass. Paret This mode is used to look for the paret of a particular fragmet. Quadrupole 2 is set o the fragmet, ad is ot scaed. The set mass that is etered i this case is the mass of the daughter (fragmet). The Start ad Ed masses specify the mass rage over which Quadrupole 1 will be scaed. The Start mass would ormally be set to a mass just below the daughter beig selected o Quadrupole 2, ad the Ed mass to a value above the highest expected paret mass. Note that there are ofte several masses from which a daughter may come, so that ay oe fragmet is derived from a umber of differet peaks. The collisio eergy is the eergy of the ios as they pass through the collisio cell. MS2 I this mode the ios are detected o the fial detector. Quadrupole 2 has both RF ad DC applied to the aalyser rods, ad is therefore resolvig. Quadrupole 1 has o resolvig DC potetials applied to the rods, ad therefore trasmits ios over a wide mass rage. The resolutio, io eergy ad io eergy ramp of MS2 will cotrol the beam shape. It has to be remembered that the io eergy (ad ramp) of Quadrupole 1 will also affect the trasmissio i this mode, as will leses 5 ad 6, which are ormally associated with Quadrupole 1. While this scaig mode ca be used for acquirig data oto disk it is mostly used i the tue widow, for settig ad optimisig the acquisitio coditios. The Start ad Ed masses specify the mass rage over which Quadrupole 2 will be scaed. Page 71

76 Q1F This mode fuctios i a similar fashio to MS2 mode from the operators poit of view. I this case however the resolvig quadrupole is Quadrupole 1, with Quadrupole 2 ot resolvig ad RF applied oly to the aalysig rods. This little used mode, is useful for fault fidig. Neutral Loss I this mode, the peak i a spectrum that will give the eutral loss specified is detected. The precursor mass is scaed i Quadrupole 1, ad Quadrupole 2 is scaed at this mass less the eutral loss mass. Startig masses are therefore detected o the mass scale of MS1. It should be remembered that the Start mass etered (for Quadrupole 1) should be greater tha the eutral loss value to give Quadrupole 2 a valid start mass. Neutral Gai This is a ifrequetly used mode, sice the mass selected by Quadrupole 2 is higher tha that of Quadrupole 1 by the eutral gai mass. It is applicable to studies where a precursor io gais mass by io molecule reactio. It also applies to fragmet ios of multiply charged ios that fragmet ito particles with a higher value of mass to charge. Figure 5.10 MS/MS editor for paret sca The fields i the MS/MS editor are maily the same as the oes i the full sca editor with the followig differeces. Mass Specifies the mass ( i.e. Daughter Mass, Paret Mass etc. ) that will be used for the particular fuctio type. A sigle mass ca be take from the Spectrum widow by selectig the appropriate mass o ay graph with a sigle right butto click of the mouse. The start ad ed masses are selected i a similar way but utilisig a right butto click-drag-release operatio. Collisio Eergy Specifies the collisio eergy i electro volts that will be used for the collisio cell durig the sca. Page 72

77 Use Tue Page Settigs Whe this cotrol is selected the collisio eergy set o the tue page is used. The collisio eergy value caot be altered by typig ew values ito tue page durig the acquisitio sice the ew values will ot be dowloaded durig the acquisitio. If you eed to alter the collisio eergy by typig ew values ito tue page durig the acquisitio you should acquire from the tue page. Collisio Eergy Ramp To apply a ramp to the collisio eergy check the Use Collisio Eergy Ramp box, this eables the CE Ramp butto. Press this butto to load the Collisio Eergy Ramp dialog. Figure 5.11 Collisio Eergy Ramp dialog Two values of collisio eergy are defied at two particular masses usig the Start ad Ed Mass cotrols ad the Collisio Eergy Start Volts ad Collisio Eergy Ed Volts cotrols. These values defie a gradiet for the collisio eergy which is the extrapolated to cover the full mass rage of the fuctio. Page 73

78 Settig up a MRM Fuctio Multiple Reactio Moitorig ( MRM ) fuctios are oly appropriate to istrumets equipped to do MS/MS. MRM fuctios are set up much i the same way as SIR fuctios but allow you to moitor a umber of MS/MS trasitios ( fragmetatios ) betwee MS1 ad MS2. Figure 5.12 MS/MS editor for MRM fuctio May of the fields i the MRM editor are the same as those i the SIR editor. There are however some differeces. Paret & Daughter Masses Specify the trasitio betwee MS1 ad MS2. A mass ca be etered either by typig its value ito the edit box at the top of the list ad pressig Eter or the Add butto, or by pullig them directly from a spectrum displayed i the Spectrum widow. To do this positio the cursor i the Paret box, the click with the right mouse butto o the io i the Spectrum widow. Repeat for the Daughter mass. The mass will be etered i the Paret ad Daughter fields of the MRM editor. Coll Eergy Specifies the collisio eergy i electro volts that will be used for the collisio cell durig the sca. Page 74

79 Q-Tof Fuctio Editig Survey Fuctios Q-Tof fuctios are set up i the same way as for other machies. The differeces are that they perform MS, MSMS ad Survey scas, ad oly ESP ad APcI io modes are available. For MS ad MSMS fuctios the Fuctio editor is the same as for other machies with the additio of oe field Scas/Spectrum. This field idicates the umber of MCA scas that are summed for each spectrum. Note: The fuctio list editor will ot add Survey fuctios to the list if o-survey fuctios are preset. If there are fuctios displayed i the Fuctio List Editor the delete them before attemptig to add a survey fuctio. A MS Survey fuctio is used to select suitable precursor ios for further MSMS aalysis. Whe the acquisitio is performed the MS Survey fuctio will be executed first. The software examies the data i each spectrum, as it is acquired, ad whe a precursor mass of iterest is detected a MSMS fuctio based o the precursor mass is geerated ad executed. As the MSMS fuctio is executed, the software examies the data i each spectrum for each fuctio, as it is acquired. If the precursor mass is o loger of iterest the executio of the MSMS fuctio will be termiated. Whe all MSMS fuctios for the curret precursor mass have bee completed the software will resume executio of the origial MS Survey fuctio, util aother precursor mass is detected whe it will execute the MSMS fuctio agai. This switchig will be repeated util the acquisitio is complete. To access the MS Survey Sca dialog press the Survey butto i the Sca Fuctios editor. Page 75

80 Figure 5.13 Survey fuctio editor MS page Survey scas are used to search for precursor ios. If automatic fuctio switchig is required check the Automatic Switchig box, otherwise maual o the fly switchig ca be carried out. Eter MS details as required. Whe the OK butto is pressed the MSMS sca dialog is displayed. Page 76

81 Figure 5.14 Survey fuctio editor MSMS page Eter the MSMS details as required ad press OK. Two fuctios will be added to the fuctio list as i Figure Figure 5.15 Fuctio List Page 77

82 Automatic Fuctio Switchig To set up Automatic Fuctio Switchig parameters select Automatic Switchig Setup from the Fuctio meu. MS to MSMS Page Figure 5.16 MS to MS/MS Switchig page The MS fuctio will stop whe the MS to MSMS Switch criteria are met. MS to MSMS Switch Criteria TIC If this optio is selected, ad the TIC of the spectrum rises above the specified Threshold. Itesity If this optio is selected, ad the itesity of the largest peak rises above the specified Threshold. Detectio Widow Whe a peak top is foud o other peaks will be looked for i the specified widow. Number of compoets It is possible to carry out automated MSMS o up to 8 coelutig precursors, eter a value betwee 1 ad 8 i this box. The umber of o coelutig precursors i a sigle ru is ot limited. Page 78

83 Charge State Tolerace Widow Do ot chage this from the default value of 3. Mass Extractio Widow The data for the masses withi this widow aroud a detected precursor is cetroided. If the itesity of the mai peak is above the (%) Itesity of Mai Peak it is selected. A mass is valid if it is ot o the exclude list, ad it satisfies the precursor selectio criteria. Precursor Selectio If Automatic is selected all valid masses satisfyig selectio criteria are moitored. If Iclude masses oly is selected oly masses i the iclude list are moitored. If Iclude masses ad Automatic is selected masses o the Iclude list will be give priority. If less tha precursors are foud the other valid masses will be moitored ( = Number of compoets). A mass is valid if it is ot o the exclude list, ad it satisfies the precursor selectio criteria. Detected Precursor Iclusio Auto exclude Do ot use this precursor to switch to MSMS agai. Always Iclude Use this precursor agai for MSMS switchig. Iclude after time Oly use this precursor to switch to MSMS whe the specified time period has elapsed after the precursor was last detected. Data Check the Discard uiterestig survey scas box to store oly the survey scas that detect precursor ios ad iitiate MS switchig. This saves o disk storage space as it does ot save survey scas which do ot cotai relevat data. Page 79

84 Collisio Eergy Page Figure 5.17 Collisio Eergy page Use Default Collisio Eergy Select this optio to use the collisio eergy defied o the MSMS Sca Fuctio page (see Figure 5.14). Use Collisio Eergy Profile Select this optio to use a collisio eergy profile. This is a file which specifies a umber of collisio eergies to use for defied masses. E.g. i Figure 5.18 if a precursor is detected i the first 100 secods ad it s mass lies betwee 50 ad 100, the perform scas at 20, 25, 30, 35 ad 40 ev util the MSMS of the precursor is termiated. To select a file press the Browse butto ad select the required file from the dialog displayed. To chage the details of a file or to create a ew oe press the Modify butto. The Modify CE Profile dialog is displayed. Page 80

85 Figure 5.18 Modify CE Profile dialog To create a ew file, press the New butto. Ay previous details are cleared. To add a profile, press the Add butto. The Collisio Eergy dialog is displayed. Figure 5.19 Collisio Eergy dialog Eter the From Mass, To Mass, From Time ad To Time required. Check the boxes for the CE 1 to CE 5 ad eter the required collisio eergy. Note: all these values are optioal. To delete a profile click o it with the left mouse butto ad press the Delete key. Page 81

86 Whe all profiles have bee defied press the Save As butto, eter the ame to save the file as ad press OK. Pressig the Save butto will save the file uder the previous ame. Use Charge State Recogitio Select this optio whe charge state recogitio is beig used to select masses. Maximum umber of Charge States Eter the umber of charge states to moitor. Rage 1 to 4. To select a Charge State file press the Browse butto ad select the required file from the dialog displayed. The file cotais a list of masses ad collisio eergies. From this list a graph is costructed which calculates the values for the masses i betwee e.g. usig Figure 5.20 for a mass of 150 a collisio eergy of 15 ev would be used. To chage the details of a file or to create a ew oe press the Modify butto. The Modify Charge State dialog is displayed. Click o the tab for the relevat page (Modify CS1, Modify CS2 etc). Figure 5.20 Modify Charge State dialog To create a ew file, press the New butto. Ay previous details are cleared. To add a profile, press the Add butto. The Charge State Mass dialog is displayed. Page 82

87 Figure 5.21 Charge State Mass dialog Eter the CS Mass ad Collisio Eergy required. To delete a etry click o it with the left mouse butto ad press the Delete key. Whe all etries have bee defied press the Save As butto, eter the ame to save the file as ad press OK. Pressig the Save butto will save the file uder the previous ame. Iclude Masses Page Figure 5.22 Iclude Masses page Masses ad rages ca be specified by checkig the Listed box ad eterig the mass or rage. Rages take the form mass1_mass2 ad ca be comma delimited i.e. 100_200,250_300. Masses ca also be specified i a text file alog with other criteria such as Retetio Time or Charge State. Page 83

88 To use a previously defied file, check the File box, press the Browse butto ad select the text file from the dialog displayed. To add a ew mass press the Add butto. This displays the Iclude Masses dialog. Figure 5.23 Iclude Masses dialog Eter the Iclude Mass required. Check the boxes for ay other values required ad eter a appropriate value. Note: All these values are optioal. E.g. If oly the mass is etered the the mass will always be icluded. If a retetio time is etered as well the the mass will oly be icluded at the specified retetio time. To delete a mass click o it with the left mouse butto ad press the Delete key. To delete all masses press the New butto. Whe all masses have bee defied press the Save As butto, eter the ame to save the file as ad press OK. Pressig the Save butto will save the file uder the previous ame. Charge State Check the Iclude By Charge State box to iclude masses by their charge state ad eter the Charge State ad the Number of Iclude Compoets. The Number of Iclude Compoets is the umber of potetial cadidate precursors based o the MS to MS criteria, the iclude ad exclude criteria ad charge state that will be used for MSMS scaig. If masses are defied i the Listed box or a File the the software checks the charge state ad if it matches the value i the Charge State box the the mass is icluded. Page 84

89 Exclude Masses Page If a mass is o the exclude list, it will ot be cosidered for detectio. Figure 5.24 Exclude Masses page Masses ad rages ca be specified by checkig the Listed box ad eterig the mass or rage. Rages take the form mass1_mass2 ad ca be comma delimited i.e. 100_200,250_300. Masses ca also be specified i a text file alog with a Retetio Time. To use a previously defied file, check the File box, press the Browse butto ad select the text file from the dialog displayed. To add a ew mass press the Add butto. This displays the Exclude Masses dialog. Figure 5.25 Exclude Masses dialog Page 85

90 Eter the Exclude Mass required. Optioally, check the Exclude Time box ad eter a appropriate value. E.g. If oly the mass is etered the the mass will always be excluded. If a exclude time is etered as well the the mass will oly be excluded at the specified time. To delete a mass click o it with the left mouse butto ad press the Delete key. To delete all masses press the New butto. Whe all masses have bee defied press the Save As butto, eter the ame to save the file as ad press OK. Pressig the Save butto will save the file uder the previous ame. Adduct Masses Page Figure 5.26 Adduct Masses page Adduct Masses ca be specified i a text file. To use a previously defied file, check the File box, press the Browse butto ad select the text file from the dialog displayed. To add a ew mass, press the Add butto. This displays the Iclude Masses dialog. Figure 5.27 Adduct Mass dialog Page 86

91 Eter the Adduct mass i the Step Size box ad press OK. To delete a mass click o it with the left mouse butto ad press the Delete key. To delete all masses press the New butto. Whe all masses have bee defied press the Save As butto, eter the ame to save the file as ad press OK. Pressig the Save butto will save the file uder the previous ame. If a adduct file is used ad a iclude file is used, all adducts will be combied with all masses o the iclude list, together with their charge states, ad the resultig masses added to the exclude list. E.g., if the iclude list cotaied the masses 100 ad 150, both with a charge state of 2, ad the adduct file cotaied the mass (Step Size) 21 The followig masses would be added to the exclude list /2, , /2, If a adduct file is specified, ad a mass is beig cosidered for switchig, its adducted masses will automatically be excluded. Page 87

92 MSMS to MS Page Figure 5.28 MSMS to MS page Whe MSMS fuctios have bee geerated, they will be carried out i parallel util the criteria specified for switchig from MSMS to MS are satisfied. If the MSMS to MS Switch method is Default, the MSMS fuctio will stop whe the MSMS to MS Switch criteria are met. If the MSMS to MS Switch method is After Time, the MSMS fuctio will stop whe the MSMS to MS Switch criteria are met, or whe the specified time has elapsed. The MSMS to MS criteria are as follows If TIC fallig below threshold is selected, ad the TIC of the spectrum falls below the specified Threshold. If TIC risig above threshold is selected, ad the TIC of all MSMS spectra for a precursor rises above the specified Threshold. If Itesity fallig below threshold is selected, ad the itesity of the largest peak falls below the specified Threshold. If the After Mass optio is selected, oly masses after the specified mass will be used i the MSMS to MS criteria. Whe all MSMS fuctios have stopped, the MS Survey fuctio will be carried out agai. Page 88

93 Other Fuctio Meu Optios If Store as table is selected, acquired fuctios will be stored as separate fuctios i the data file. If ot selected, MSMS scas will be stored as oe fuctio i the same data file if they are ruig cosecutively, ad i cosecutive data file fuctios if they are ruig i parallel. If Costruct from spectrum is switched o, a right mouse butto click ad drag operatio o a previously acquired spectrum will eter the Start ad Ed masses i the fuctio editor. For MSMS fuctios a sigle right mouse click will eter the mass i the MSMS of field. Note: Q-Tof ca have up to 32 MS ad MSMS fuctios, but oly oe Survey fuctio. Moitorig Acquisitios Figure 5.29 Automatic Fuctio switchig status dialog Whe a acquisitio is started the automatic switchig status dialog is displayed, it shows the precursors curretly ruig. Pressig the Stop butto will stop moitorig the curret precursor. Pressig the Stop All butto will stop moitorig all precursors. Page 89

94 Figure 5.30 Maual o the fly Fuctio switchig status dialog If Automatic Switchig was ot selected, whe a acquisitio is started the Maual o the fly switchig status dialog is displayed, it shows the precursors curretly ruig. Eterig a Paret Mass ad pressig the Add butto will add a precursor to the list. Selectig a precursor from the list ad pressig the Delete butto will delete the precursor from the list. Pressig the Delete All butto delete all precursors. Selectig a precursor from the list ad pressig the Load butto will geerate a MSMS fuctio ad sed it to the istrumet. Pressig the Load All butto will do this for all precursors. Pressig the Stop butto will stop moitorig the curret precursor. Pressig the Stop All butto will stop moitorig all precursors. Page 90

95 Notes Page 91

96 Notes Page 92

97 Acquirig Data Startig a Acquisitio Chapter 6 Sigle Sample There are two ways of startig a acquisitio, a sigle sample acquisitio from the Tue page or a multiple sample oe from the MassLyx Top level scree. To start a sigle sample acquisitio 1. Press the Acquire butto o the Tue page or choose Acquire from the Tue page Widow meu. 2. Eter required data. 3. Press the Start Acquisitio butto. Figure 6.1 Start acquisitio dialog Page 93

98 Multiple Samples The MassLyx top level scree cotais a Sample List Editor for defiig multiple samples which may be used together to perform a quatitative aalysis. The list of samples is set up usig a spreadsheet style editor, which ca be tailored to suit differet requiremets. To start a multi sample acquisitio 1. Set up a Sample List (see MassLyx NT User Guide, Sample Lists for details). 2. Choose Start from the top level Ru meu or press the toolbar butto. This displays the Start Sample List Ru dialog. 3. Check the Acquire Sample Data, Auto Process Samples ad Auto Quatify Samples boxes as required. 4. Eter values i the Ru From Sample ad To Sample boxes. The default is all samples i the list. 5. Check the Priority ad/or Night Time Process boxes as required. See the Gettig Started chapter of the MassLyx maual for details. 6. Press the OK butto. 7. Repeat steps 1 to 5 as required. Sample Lists will be added to a queue ad ru sequetially uless Priority or Night Time Process has bee checked. Figure 6.2 Start Sample List Ru dialog 8. The sample, i the Sample List, which is curretly beig acquired will have a ext to it. Page 94

99 Process The Process cotrols allow you ru processes before ad after the acquisitio. Use the Pre-Ru cotrol to specify the ame of a process that will be ru before acquisitio of the files i the sample list. Use the Post-Ru cotrol to specify the ame of a process which will be ru after acquisitio of the files i the sample list, this could be used for example, to switch the istrumet out of operate ad to switch off various gases. If you wish to ru a process after each sample i the sample list has bee acquired, format the Sample List to display the Process colum ad eter the ame of the process to be ru for each of the samples. If you wish the process to automatically operate o the data file, which has just bee acquired, the the Use last acquired file as default parameter i the MassLyx System Globals dialog should be left uchecked. The MassLyx System Globals dialog is accessed by choosig System Globals from the MassLyx Customize meu. Page 95

100 Automated Aalysis of Sample List Select Process Samples from the Quatify meu to display the Quatify Samples dialog. Check the boxes required ad press OK. Figure 6.3 Quatify Samples dialog The Quatify Samples dialog allows you to automatically process data files oce they have bee acquired. To perform Itegratio, calibratio of stadards, quatificatio of samples ad pritig of quatificatio reports select the relevat check boxes. See the Quatify Chapter, MassLyx User Guide, for more detailed iformatio about usig automated sample list aalysis. Itegrate Samples Itegrates all the sample data files amed i the Peak List. Calibrate Stadards Uses Itegratio results to form Quatify calibratio curves. Quatify Samples Uses Itegratio results ad Quatify calibratio curves to calculate compoud cocetratios. Prit Quatify Reports Produces hard copies of the results of itegratio ad quatificatio. Export Results to LIMS Produces a text file cotaiig the quatificatio results details for use with LIMS systems. If this box is checked the LIMS Export File Browse butto becomes eabled, press the Browse butto, select a file or eter the ame of a ew oe ad press Save. Project The Project field displays the project ito which data will be acquired. If you wish to chage the project ito which data will be acquired, you ca cacel the acquisitio ad create a ew project by choosig Project Wizard, or ope a existig oe by choosig Ope Project, from the MassLyx Top Level File meu. Aalyse From Sample To Sample Sets the rage of samples i the sample list which will be aalysed. Page 96

101 Moitorig a Acquisitio Acquisitio status is also show o the MassLyx scree. The ru time is show o the MS pael ad the sca status, sample umber ad sca umber are show o the Status bar at the bottom of the page. The Acquisitio Status Widow The acquisitio status widow provides a sca by sca statistical report of the progress of a acquisitio. The MCA Display Figure 6.4 Acquisitio Status dialog If you are acquirig MCA data you ca watch the data accumulatig i real time usig the MCA display. This display shows a spectrum as well as some statistical iformatio. Figure 6.5 Real time MCA Display dialog Page 97

102 Chromatogram Real-time Update To view the chromatogram that is curretly beig acquired i real time 1. Ope the data file usig the MassLyx data browser. 2. Press the toolbar butto or select Real-Time Update from the Display meu. The chromatogram display will be updated as the acquisitio proceeds. Spectrum Real-time Update To view the spectrum that is curretly beig acquired i real time 1. Ope the data file usig the MassLyx data browser. 2. Press the toolbar butto or select Real-Time Update from the Display meu. Figure 6.6 Spectrum Real-Time Update dialog To tur real time update o select the Eable Real-Time update check box, to tur it off clear this check box. Whe real-time update is o the display will cotiually be updated with spectra from the curret acquisitio. The actual iformatio displayed is determied by selectig oe of the followig radio buttos. Real-time update ca also be tured o ad off via the Real-Time spectrum toolbar butto. Selectig the Latest sca butto will display the last acquired sca, this is the default optio. Selectig the Average all scas will update the display with spectra formed by averagig all the spectra that have so far bee acquired. Selectig the Average latest scas butto will update the display with spectra formed by averagig the last scas acquired, where is specified i the associated edit cotrol. Page 98

103 Stoppig a Acquisitio From the Tue Page, press the Stop butto. From the MassLyx scree choose Stop from the Ru meu or press the toolbar butto. Data acquired up to this poit will be saved. Page 99

104 Automatic Startup ad Shutdow MassLyx comes with automatic Startup ad Shutdow files. They are foud i the C:\Masslyx\Shutdow directory ad are called ShutDowxxx.acl ad StartUpxxx.acl where xxx refers to the istrumet cofiguratio. E.g. ShutDowESI_ACE.acl for a istrumet cofigured as a GC or LC system (ACE). Whe Startup or Shutdow is selected from the MassLyx Ru meu it is these files which are ru. The Shutdow Editor The shutdow editor allows the automatic startup ad shutdow procedures to be modified or ew procedures to be created. To access the editor select Edit Shutdow from the MassLyx Ru meu. Figure 6.7 Shutdow editor Check the Eable Startup before batch box to perform the startup tasks whe a Sample List is submitted. Check the Eable Shutdow after batch box to perform the shutdow tasks after a batch of samples has completed. Eter a time i the Shutdow miutes after batch or error box at which to perform the shutdow tasks. There is a optio to perform the shutdow tasks immediately after a error occurs or after the time defied i the Shutdow miutes after batch or error box. If the Eable Shutdow o error box is checked the the shutdow tasks are performed after the defied time. Note the Eable immediate Shutdow o error box is grayed out if this optio is selected. If the Eable immediate Shutdow o error box is checked the the shutdow tasks are performed as soo as the error is detected. Note the Eable Shutdow o error box is grayed out if this optio is selected, but the shutdow time ca still be chaged as this applies to the Eable Shutdow after batch optio. Page 100

105 The Auto Cotrol Tasks Page Figure 6.8 Auto Cotrol Tasks editor Task This is a dropdow list box with all the available tasks. Pre-Delay This is the legth of time that will elapse before the curret task is performed. Post-Delay This is the legth of time that will elapse after the curret task has bee completed ad before the ext task is started. E.g. a Post delay of 60s, i the Vet Istrumet task i Figure 6.8, meas that there will be a delay of 60 secods before the ext task is started, to allow the machie to vet fully. Io Mode This is a dropdow list box with all the available Ioisatio Modes. File Name This is the ame of the Tue file to be used. The file ame ca be typed i, icludig the full path ame, or selected from the browser displayed whe the butto is pressed. To Add a Task 1. Select a task from the dropdow Task list box. 2. Eter the required parameters. 3. Press the add butto. Note. If this is a ew task timetable the task will be added to the ed of the list. If a task has bee iserted ito the task timetable the all subsequet tasks will be added after the iserted task. To add a task to the ed of the timetable after isertig a task, click twice with the left mouse butto below the last etry i the timetable ad the add the ew task. Page 101

106 To Isert a Task 1. Click, with the left mouse butto, o the etry i the task timetable before which you wat to isert the ew task. 2. Select a task from the dropdow Task list box. 3. Eter the required parameters. 4. Press the add butto. The task will be iserted before the selected etry. To Modify a Task 1. Click, with the left mouse butto, o the etry i the task timetable. The details for the task will be displayed i the fields o the left of the scree. 2. Chage the required parameters. 3. Press the modify butto. The details will chage i the task timetable. To Delete a Task 1. Click, with the left mouse butto, o the etry i the task timetable. The details for the task will be displayed i the fields o the left of the scree. 2. Press the add butto. The task selected will be deleted from the task timetable. To Delete All Tasks Press the add butto. All tasks will be deleted from the task timetable. To chage the width of a colum The width of the colums ca be chaged, by positioig the mouse poiter o the headig betwee two colums util the symbol appears, ad the clickig the left mouse butto ad draggig util the colum is the required width. Page 102

107 The Shutdow Editor Toolbar Toolbar butto Meu equivalet File New File Ope File Save or File Save As File Prit Purpose Create a ew Startup or Shutdow file Ope a existig Startup or Shutdow file Save a Startup or Shutdow file Prit a Startup or Shutdow file Cotrol List Ru List Ru a Startup or Shutdow file Cotrol List Stop List Stop a Startup or Shutdow file Help Help Topics Ivoke help Page 103

108 Savig/Loadig Startup ad Shutdow Files To Ope a Startup or Shutdow file 1. Press the Toolbar butto or select Ope from the File meu. This displays the Ope file dialog. Figure 6.9 The Ope File dialog 2. Select a data file ad press the Ope butto. To Save a Startup or Shutdow file 1. Press the Toolbar butto or select Save or Save As from the File meu. If this is a ew file, or the Save As optio has bee selected, the Save As dialog is displayed Figure 6.10 The Save File dialog 2. Type a ame ito the File Name box ad press the Save butto. Page 104

109 Pritig Startup ad Shutdow Files To Prit a Startup or Shutdow File 1. Press the Toolbar butto or select Prit from the File meu. This displays the Prit dialog. Figure 6.11 The Prit dialog 2. Select the priter, prit rage ad umber of copies ad press the OK butto. Creatig Startup ad Shutdow Files To Create a Startup or Shutdow File 1. Press the butto or select New from the File meu. Ruig Startup ad Shutdow Files If Startup or Shutdow is select from the MassLyx Ru meu or from the Shutdow editor Cotrol List meu the the automatic startup ad shutdow files are ru. If you wish to ru a differet Startup or shutdow file. Ope the required file i the Shutdow editor ad press the toolbar butto or select Ru List from the Shutdow editor Cotrol List meu. Press the toolbar butto or select Stop List from the Shutdow editor Cotrol List meu if you wish to stop ruig this file. Page 105