3500 series software...the world leader in serving science 3500 series instrument data collection...
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The world leader in serving science
3500 Series Instrument Data Collection Software Version 4.0 and GeneMapperTM ID-X Software v1.6Dr. Anke KrugerManager HID Field Application Specialists Europe
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Data Collection Software v4 enhancements:• Signal optimization • Pull-up reduction• Offscale recovery• Updated Library Organization• Flexible plate loading• Additional features
GeneMapper ID-X v1.6:• Off-scale Data Recovery• Export Options for Probabilistc Genotyping• Peak Detection Algorithm changed• Enhanced Plot and Printing Options• Sorting Improvements
Agenda
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3500 Series Instrument Data Collection Software Version 4.0
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Computer Requirements
Operating System
• Win 10 2016 LTSB, 64bit• Java 7 32bit• Tomcat 7.0.82 32bit• Verification testing included: Symantec Endpoint Protection 12 and McAfee
Endpoint Security 10.5 and Windows Defender. • McAfee Endpoint Security may block services startup. See release notes for
details.
Dell OptiPlex XE2 Computer
Updates
• Server Monitor updated • Patch for camera corrupt error• No licensing• FSE installed
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Signal optimization
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• Goal: reduce capillary-to-capillary variation within an injection• Spatial approach developed
Signal optimization
• Utilize spatial calibration data• 24-capillary instruments only• Automatically applied
Optical approach: Reduce variation introduced by optics
Optimize Signal
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• Displayed in Spatial Calibration screen• Included in the Spatial calibration report, can be exported or
printed• 8 capillary instruments: factor = 1 for all capillaries
Signal Optimization Factor
Reduce Variation Due to Optics
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Signal Optimization Results
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Pull-up Reduction
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Recognizing Pull-up
• Analyzed data: centered under parent peak
• Raw data: same shape as parent peak
Spectral
• Analyzed data: not centered under parent peak
• Raw data: sinusoidal curve
Instrument-Specific
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Pull-Up Reduction in DC v4.0
Pull-up reduction
Addresses spectral (including off-scale)
and instrument-specific pull-up
Utilizes sample-specific spectral data
To enable pull-up reduction
Utilize an Applied Biosystems (AB) STR
kit
Perform spectral calibration using an
AB dye matrix and AB dye set
Choose applicable AB assay
Spectral correction
Takes place prior to creation of .hid files
Raw Data viewed in GeneMapper IDX (all versions) will have the
reduction applied
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Pull-Up Reduction in DC v4.0
• Identified by labeling as allelic ladder sample type
• Identified by density of peaks in profile
Allelic ladders – pull-up reduction DISABLED
• Signal-to-noise ratio
• Number of peaks and dyes detected
• Calibrated matrix will still be applied (i.e. sample-specific spectral data not utilized due to poor quality)
Poor quality samples – pull-up reduction DISABLED
• Adds approximately 2-3 minutes per injection for processing (3500xL)
• Pull-up frequency is reduced
• Peak height percentage of remaining pull-up generally falls within 1-3%
Important Notes
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Improved Data Interpretation – Pull-up
Single-source 1ng sample amplified with the GlobalFiler kit analyzed at 50 RFU31 pull-up edits
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Improved Data Interpretation – Pull-up Reduction
The same GlobalFiler profile analyzed at 50 RFU after pull-up reduction is applied.6 pull-up edits
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Off-scale data recovery
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• Capture “off-scale”• Truer peak height• Decrease manual interpretation
• Off-scale data now approaching ~65,000 RFU
• Global filters more effective• Stutter filters more applicable
• Decrease pull-up associated with off-scale peak
• On-scale data not impacted
• Higher 1st pass success rate
Off-Scale Data Recovery (OSR) for Database Samples
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Off-Scale Data Recovery
• Bin data signals saturation• Bin data converted to dye data• Dye data capped at ~32K RFU
values
Saturation
• CCD camera saturation limit is extended (OSR assays)
• More bin data can be utilized• Dye data capped at ~65K RFU
values
Off-Scale Data Recovery
GMIDX 1.6 is required to analyze/view OSR data above ~32K RFU.
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Off-Scale Data Recovery Field Testing Results
OSR Disabled(Any6Dye dye set)
OSR Enabled(J6 dye set)
Test Site 1 5 markers in 2 samples 0 markers in 0 samplesTest Site 3 99 markers in 12 samples 4 markers in 1 sample
Test Site 2 5 markers in 2 samples 0 markers in 0 samples
(10) Reference GFE samples(3) GFE Positive Controls
Data from Test Site 1
OSR ON
OSR OFF
Using OSR dye set enables OSR feature – works with AB dyes
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Additional Features
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Flexible Plate Loading - 3500 Series
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Added Flexibility and Control – System Preferences
• Instrument settings
• Uncheck “allow runs” options
• Access: admins only
• Results in pre-run validation check error
Choose: hard stop
Admin view
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DC 4.0 Enhancements - Summary
Improved data analysis options• Pull-Up Reduction• Off-Scale Data Recovery (OSR)• Signal Optimization (SO)
Improved user experience• No yearly licensing• Flexible Plate Loading• 6-dye HID Install Standard• Library Assets Updated• Flexible admin preferences for hard stops and warnings• Bi-weekly scheduling• Exportable consumables list• Exportable injection list• Normalization Factor and average peak height in Sample View• EPT for completed/aborted runs
User Bulletin available
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GeneMapperTM ID-X Software v1.6
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GMIDX updates including:
• Operating Systems (Win 10, 64bit and Win7, 64 bit compatibilty)
• Off-scale Data Recovery
• Export Options for Probabilistc Genotyping
• Peak Detection Algorithm changed
• Command Line Interface
• Profile Comparsion Tool
• Enhanced Plot and Printing Options
• Sorting Improvements
• Panel and Report Manager Updates
• Support of SeqStudio file format
• ...
Agenda
HID validation expected H2 of 2019
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Off-Scale Data Recovery
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Off-Scale Data Recovery
• Upper limit: ~65,000 RFU
• MUST use GeneMapper™ IDX v 1.6
RFU upper limit extended for DC 4.0
• Global filters more effective
• Stutter filters more applicable
• Decreased pull up
• Higher 1st pass success rate
• On-scale data not impacted
Better peak height estimate
• Peaks will be capped
• Secondary dye colors may be pulled-up to 65K
• Analyze with caution!
Severely off-scale data (>65K RFU)
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Off-Scale Data Recovery (OSR)
41251
29620
v1.6 = Pass
v1.5 = Flagged
OS w/ pink bar
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Export for Probabilistic Genotyping
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Probabilistic Genotyping – Data Export
• Two export file type options:• .txt or .csv • Select from the right-hand drop-
down menu
• When you export without stutter:• Labeled allele peaks are identified
as Allele 1 and Allele 2
• When you export with stutter:• Stutter peaks and labeled allele
peaks are labeled sequentially
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Enhanced Plot and Printing Options
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Printing
Custom Header
Marker header outlined in color
Bin Color adjusted to improve visibility
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Sorting within the Software
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Sorting Within the Software
• Projects sorted by default with most recently created on top
• Analysis Methods and Size Standard drop downs in project window are alphabetical
• Genotype Table can be sorted by Marker only without dye consideration
General Sorting
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Comparison NGM Detect - NGMSelect
Different Samples - One Marker – Different dyes
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Comparison NGM Detect - NGMSelect
Different Samples - One Marker - sample order by selection
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GMIDX v1.6 Details- Summary
Improved data analysis options• Off-Scale Data Recovery (OSR)• Data export for stutter• Peak height detection
Improved user experience• Profile Comparison Tool• CLI• Sorting (projects, drop-down menus, Genotypes Table)• Plots (scrolling, editing/deleting, size standard display, PLI, tables)• Panel manger (v6x, IQS and Ymarker functionality)• Report manger• SS Normalization• Printing (comment, marker header)• SAE: Audit record deletion, log-in list• Seq Studio file support
IT compliance• Win 7 or Win 10• CODIS updates• Chinese Language• Oracle Security
User Bulletin available
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Thank you for your time & attention
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