Applied Biosystems 7900HT Fast Real-Time PCR System and SDS ...

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Chapter 2Getting StartedGetting StartedBefore You BeginProcedureQuick ReferenceIf this is your first time using the Applied Biosystems 7900HT Fast Real-Time PCRSystem, consider completing the “Basic Software Skills Tutorial” on page 2-10before continuing. The tutorial will provide you with the fundamental knowledgerequired to operate the SDS software and will teach you timesaving techniques toallow you to use it quickly and efficiently.The following table contains a list of major procedures described inside this manual.Setting Up and Running SDS ExperimentsCreating an SDS Plate Document . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .3-9Running an Individual SDS Plate Document . . . . . . . . . . . . . . . . . . . . . . . . . . . .4-23Running Batches of SDS Plate Document . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .4-31Stopping a Run from the SDS Software . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .4-28Stopping a Run from the Automation Controller Software . . . . . . . . . . . . . . . . .4-41Ejecting a Plate from the SDS Software . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .4-29Ejecting a Plate from the Automation Controller Software . . . . . . . . . . . . . . . . .4-41Analyzing Run DataAnalyzing an Allelic Discrimination Run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .5-5Analyzing an Absolute Quantification Run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .6-5Analyzing an Relative Quantification Run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .6-15Analyzing a Dissociation Curve (Melting Curve) Run . . . . . . . . . . . . . . . . . . . . .6-37Maintaining the 7900HT InstrumentChanging the Plate Adapter . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .7-12Replacing the Sample Block . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .7-6Decontaminating the Sample Block . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .7-14Performing a Background Run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .7-16Performing a Pure Dye Run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .7-20Adding Custom Dyes to the Pure Dye Set . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .7-27Verifying Instrument Performance Using a TaqMan ® RNase P InstrumentVerification Plate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .7-30Maintaining the Automation AccessoryAdjusting the Sensitivity of the Plate Sensor Switch . . . . . . . . . . . . . . . . . . . . . .7-37Aligning the Plate Handler . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .7-41Aligning the Fixed-Position Bar Code Reader . . . . . . . . . . . . . . . . . . . . . . . . . . .7-49Cleaning and Replacing Gripper Finger Pads . . . . . . . . . . . . . . . . . . . . . . . . . . . .7-52Note: The Automation Accessory includes the Zymark ® Twister Microplate Handlerand the fixed-position bar code reader. See “Instrument Components” on page 1-3for more information.2-2 Applied Biosystems 7900HT Fast Real-Time PCR System and SDS Enterprise Database User GuideDRAFTSeptember 1, 2004 11:38 am, CH_GetStart.fm
Getting StartedUsing the SDSSoftwareOnline HelpThe Sequence Detection Systems Software Online Help can guide you through theprocedures for setting up, performing, and analyzing runs. To get help at any time,click (Help button) located inside the dialog box or window in which you areworking.The SDS software provides two ways to access the online help:To…access general helpget help for using a specific dialog box,plot, or featureThen…select Help > SDS Online Help.click (help button) located inside the dialogbox or window in which you are working.For MoreInformationFor information about the Applied Biosystems 7900HT Fast Real-Time PCR Systemor the SDS software, Applied Biosystems recommends the following references:• Applied Biosystems 7900HT Fast Real-Time PCR System Site PreparationGuide (PN 4317595)• Applied Biosystems 7900HT Fast Real-Time PCR System Quick Starts for:– Allelic Discrimination (PN 4351673)– Absolute Quantification (PN 4351672)– Relative Quantification (PN 4351674)• Sequence Detection Systems Software Online Help• Microsoft Windows Operating System Online Help• SDS Enterprise Database for the Applied Biosystems 7900HT Fast Real-TimePCR System Administrators Guide (PN 4351669)Applied Biosystems 7900HT Fast Real-Time PCR System and SDS Enterprise Database User Guide 2-3DRAFTSeptember 1, 2004 11:38 am, CH_GetStart.fm
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Applied Biosystems7900HT Fast Real-
Page 3 and 4: ContentsPrefaceHow to Use This Guid
Page 5 and 6: Chapter 4Operating the InstrumentNo
Page 7 and 8: Chapter 7Maintaining the Instrument
Page 9 and 10: Appendix D Theory of OperationFluor
Page 11 and 12: PrefaceHow to Use This GuidePurpose
Page 13 and 14: Safety and EMC Compliance Informati
Page 15 and 16: Symbols on InstrumentsSymbols on In
Page 17 and 18: General Instrument SafetyGeneral In
Page 19 and 20: Chemical Waste SafetyChemical Safet
Page 21 and 22: Physical Hazard SafetyOvervoltageRa
Page 23 and 24: Workstation SafetyWorkstation Safet
Page 25 and 26: Product Overview 11In This Chapter
Page 27 and 28: 7900HT FAST Real-Time PCR SystemSec
Page 29 and 30: 7900HT InstrumentInterchangeableThe
Page 31 and 32: 7900HT FAST Real-Time PCR SystemBar
Page 33 and 34: Compatible ConsumablesCompatible Co
Page 35 and 36: Instrument ConnectionsFixed-Positio
Page 37 and 38: Section 1.2 Getting to Know the Sof
Page 39 and 40: SDS Software Related Files and Form
Page 41 and 42: 7900HT FAST Real-Time PCR SystemMan
Page 43 and 44: Esc F1 F2 F3 F4 F5 F6 F7 F8 F9 F10
Page 45 and 46: Section 1.3 SDS Enterprise Database
Page 47 and 48: * + Enter* +Enter3Down UpNumLockSys
Page 49 and 50: About the SDS Enterprise Database S
Page 51 and 52: Esc F1 F2 F3 F4 F5 F6 F7 F8 F9 F10
Page 53: Getting Started 22In This Chapter G
Page 57 and 58: GR23967900HTPowering On the 7900HT
Page 59 and 60: Using the SDS SoftwareUsing the SDS
Page 61 and 62: Using the SDS SoftwareAbout theMess
Page 63 and 64: Basic Software Skills TutorialNotes
Page 65 and 66: Basic Software Skills TutorialExerc
Page 67 and 68: Basic Software Skills TutorialExpor
Page 73 and 74: Basic Software Skills TutorialLesso
Page 77 and 78: Basic Software Skills Tutorial3. Se
Page 81 and 82: Using SDS Plate DocumentsUsing SDS
Page 83 and 84: Using SDS Plate DocumentsWell Inspe
Page 85 and 86: Preparing a Run 33In This Chapter N
Page 87 and 88: Notes for Database UsersDescription
Page 89 and 90: Workflow OverviewWorkflow OverviewE
Page 91 and 92: Workflow OverviewAllelicDiscriminat
Page 93 and 94: Step 1 - Creating a Plate DocumentS
Page 95 and 96: Step 2 - Applying Detectors and Mar
Page 101 and 102: Step 3 - Configuring the Plate Docu
Page 103 and 104: Step 5 - Programming the Plate Docu
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Step 5 - Programming the Plate Docu
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Step 5 - Programming the Plate Docu
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Step 6 - Saving the Plate Document
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Step 7 - Creating a Plate Document
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Step 9 - Running the Plate on the 7
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Operating the Instrument 44In This
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Notes for Database UsersDescription
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Section 4.1 Consumable PreparationS
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Compatible ConsumablesTable 4-1Comp
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Preparing Optical Plates for Usea.
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Preparing TaqMan Low Density Arrays
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Section 4.2 Running an Individual P
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Running a Single Plate (Using the S
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Running a Single Plate (Using the S
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After the RunAfter the RunUser Acce
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Section 4.3 Automated OperationSect
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Operating the Software with an SDS
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Operating the Software without an S
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Page 153 and 154:
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Running Plates Using the Automation
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Running Plates Using the Automation
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Analyzing End-Point Data 55In This
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Notes for Database UsersNotes for D
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Section 5.1 Allelic DiscriminationS
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OverviewTable 5-1Signal and Sequenc
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Before You BeginBefore You BeginUsi
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Opening the Run DataOpening the Run
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Calling and Scrutinizing Allelic Di
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Page 175 and 176:
Page 177 and 178:
After the AnalysisSaving theResults
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Analyzing Real-Time Data 66In This
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Page 183 and 184:
Section 6.1 Absolute Quantification
Page 185 and 186:
OverviewAlgorithmicManipulation ofR
Page 187 and 188:
Analysis ChecklistAnalysis Checklis
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Analyzing the DataAnalyzing the Dat
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Viewing ResultsViewing ResultsViewi
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Section 6.2 Relative Quantification
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OverviewAlgorithmic Manipulation of
Page 197 and 198:
OverviewAutomatic Outlier RemovalAb
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Getting StartedGetting StartedUsing
Page 201 and 202:
Options for Analyzing Relative Quan
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Creating the Studyd. In the Plate Q
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Analyzing the Study• Manual Ct -
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Analyzing the StudySpecifying the A
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Viewing ResultsViewing ResultsDispl
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Viewing ResultsAbout DownArrowsDown
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After the AnalysisAfter the Analysi
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Section 6.3 Dissociation Curve Anal
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Before You BeginExample ResultsFigu
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Opening the Run DataOpening the Run
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Determining T m Values for the Anal
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Section 6.4 Procedure ReferenceSect
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Setting the Baseline and Threshold
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Eliminating Outlying AmplificationE
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Eliminating Outlying Amplification6
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After the AnalysisSaving theResults
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Maintaining the Instrument 77In Thi
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Page 237 and 238:
Section 7.1 Maintaining the 7900HT
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Replacing the Sample BlockMaterials
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Replacing the Sample Block9. Loosen
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7900HT FAST Real-Time PCR SystemRep
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Changing the Plate Adapter4. Place
Page 247 and 248:
Decontaminating the Sample BlockCle
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Performing a Background RunPreparin
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Performing a Background RunAnalyzin
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Performing a Pure Dye RunAfter a ru
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Performing a Pure Dye Run• Fast 9
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Performing a Pure Dye RunAnalyzing
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Adding Custom Dyes to the Pure Dye
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Adding Custom Dyes to the Pure Dye
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Verifying Instrument PerformanceTab
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Verifying Instrument PerformancePre
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Section 7.2 Maintaining the Plate H
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Adjusting the Sensitivity of the Pl
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Adjusting the Sensitivity of the Pl
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7900HT FAST Real-Time PCR SystemAli
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Aligning the Plate Handler5. Using
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Aligning the Plate Handler7. Using
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Aligning the Plate Handler13. Click
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7900HT FAST Real-Time PCR SystemAli
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Aligning the Fixed-Position Bar Cod
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Section 7.3 Maintaining the Compute
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Maintaining the SDS softwareSeveral
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Troubleshooting 88In This Chapter T
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Troubleshooting TableTable 8-1Troub
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Low Precision or IrreproducibilityL
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Low Precision or IrreproducibilityD
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Background RunsBackground RunsBackg
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Pure Dye Runs6. Repeat step 4 until
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End-Point Runs (Allelic Discriminat
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Software and 7900HT InstrumentTable
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Zymark Twister Microplate Handler a
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TaqMan Low Density ArrayTable 8-6Tr
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Software ReferenceAAIn This Appendi
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Importing Plate Document Setup Tabl
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Setup Table Files1. File VersionDes
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Setup Table Files6. Detectors ListD
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Setup Table Files9. Markers List (A
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Setup Table FilesFormat(for a singl
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Setup Table FilesFormat(for a singl
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Setup Table FilesExample Code A-16A
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Exporting Plate Document Data4. In
Page 327 and 328:
Operating the SDS Software from a C
Page 329 and 330:
Using the Search ToolLogical Operat
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Connecting SDS Software to the Data
Page 333 and 334:
Designing TaqMan Reagent-BasedAssay
Page 335 and 336:
Assay Development Guidelines• For
Page 337 and 338:
Design Tips for Allelic Discriminat
Page 339 and 340:
Kits, Reagents, and ConsumablesCCIn
Page 341 and 342:
Consumables and DisposablesConsumab
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Instrument Maintenance and Verifica
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Custom Oligonucleotide SynthesisTab
Page 347 and 348:
Theory of OperationDDIn This Append
Page 349 and 350:
Fluorescent-Based ChemistriesBasics
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Real-Time Data AnalysisPhase 2: Lin
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Real-Time Data AnalysisSignificance
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A similar equation for the endogeno
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Limited Warranty StatementEEWarrant
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BibliographyGeneral Paperson TaqMan
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Klein, D., Janda, P., Steinborn, R.
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Dolken, L., F. , Schuler, and G. ,
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AllelicDiscriminationVirtanen M., H
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Glossary.sdd.sdm.sds.sdt∆C T∆
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ackgroundbaselinecalibratorcDNA(com
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minor groovebinder (MGB)multicompon
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R nrunsessionsingle nucleotidepolym
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IndexSymbols- 6-31↑ 6-33Numerics7
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fluorescent, common sources 8-7isol
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adjusting step parameters 3-21confi
Page 381 and 382:
sample block locking bar 7-8sample
Page 384:
Worldwide Sales and SupportApplied
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