README

 

QuantStudio™ 12K Flex Software v1.2.2

For QuantStudio™ 12K Flex Real-Time PCR System

 

 

CONTENTS

OVERVIEW

FEATURES

UPDATES

LICENSE

COMPATIBILITY

KNOWN ISSUES

SYSTEM REQUIREMENTS

INSTALL SOFTWARE

INSTALLATION OF NEW MODULES

UNINSTALL SOFTWARE

 

 

OVERVIEW

QuantStudio™ 12K Flex Software v1.2.2 supports the QuantStudio™ 12K Flex Real-Time PCR System. The QuantStudio™ 12K Flex Software allows the user to open and analyze experiments generated from the QuantStudio™ 12K Flex Real-Time PCR System. The software also enables the user to set up experiments, send experiments to the instrument, control the thermal cycling process in the instrument, collect data and analyze the collected data, all in an integrated and streamlined fashion.

 

 

FEATURES

·         QuantStudio™ 12K System home screen enables launch of analysis applications from within application

·         New Sample Tracker software for OpenArray® plates

·         Easy to set up experiments for both novice and advanced users for 384-well and OpenArray® blocks

·         Customizable optical filter selection for more complex applications, such as multiplexing

·         QuickStart setup so you can begin a run immediately and enter plate information later

·         Ability to run up to four OpenArray® plate experiments at once

·         Real-time monitoring of amplification curves enables you to view a run progress (can be viewed from a remote PC as well)

·         Auto-baseline and auto-threshold for simplified data analysis

·         Alternative method for data analysis using a non-threshold CT calling method

·         Multiple Plots view for simultaneously assessing data from four perspectives

·         Automated SNP genotype calling with intuitive graphical output and quality-value assignment

·         Troubleshooting flags to help you diagnose and solve problematic experiments

·         Tool tips for easy identification of sample wells when viewing amplification curves or SNP genotyping plots

·         Email notifications to alert you when a run has started or ended

·         Easy cut-and-paste functionality

·         Export easily to Microsoft® PowerPoint® software, Excel® software, or directly as a .JPEG file

 

 

UPDATES

v1.2.2

·         Algorithm update to improve OpenArray® chip-finding 

·         Update operating temperature range setting for LED

Bug Fixes:

·         Fix incorrect parameter and display issue in Define, Assign (including Plate Layout) and Run Method page

·         Fix issue on order of Target Name in Export Result tab and files

 

v1.2.1

·         Algorithm update to enhance OpenArray® chip-finding

·         Improve RNase P pass/fail criteria for OpenArray® experiment

Bug Fixes:

·         Fix issue on OpenArray® real-time amplification plot freezing during run

·         Fix files downloading issue upon completion of run

 

v1.2

·         Allow assay specific cycle calling for genotyping experiment

·         Enhance RNase P pass/fail criteria for OpenArray® experiment

·         Show VIC® amplication plots for OpenArray® genotyping experiment

·         Show Amplification Score (Amp Score) for VIC® plots

·         Additional CqCONF flag in Analysis Settings, well table, export table and printed report

·         Allow export of ROX™ values for OpenArray® experiment

·         Allow analyzes experiment file (.EDS) from raw data in the event the software shows missing cycle data in the Amplification Plot

·         Allow download of RNase P calibration file before saving

·         Include sample name in the Print Report plate layout view

·         Extend calibrations expiration dates to one year

·         Two targets are displayed when only one target is selected  in OpenArray® dual reporter amplification plot (for genotyping)

·         Ability to save OpenArray® .EDS file after performing a calibration override

·         e-Signature features enabled for Open Array® experiment run

·         Enable command line export to ensure data for  all wells of an OpenArray® experiment are exported correctly via command line

Bug Fixes:

·         Fix incorrect rounding for Amp Score values in Export Result and CRT/CT values in well table

·         Fix the software to prevent user from closing the calibration run window when user clicks “Close”

·         Fix incorrect color patterns swapping from 96-well to 384-well for Genotyping – 16 Adjustable Pipette format in OpenArray® Sample Tracker Software

·         Fix incorrect mapping from 96-well to OpenArray® for Genotyping – 16 Adjustable Pipette format in OpenArray® Sample Tracker Software

·         Firmware update to include a thermal protocol to clean the OpenArray® block in the instrument

 

v1.1.2

·         Algorithm update to reduce false CRT calls of Undetermined

·         Firmware update with algorithm enhancement to improve sensitivity of chip detection

 

v1.1.1

·         Algorithm updates for system stability

·         Defect repair during automation run

·         Defect repair in Sample Tracker Software

·         Defect repair for import of .TPF file when using 196-format and 168-format for OpenArray® experiments

 

 

LICENSE

See accompanied End User License Agreement for details. You must agree to the terms of the license before installing or using the software.

                            

 

COMPATIBILITY

·         The software can open experiment files created with QuantStudio™ 12K Flex Software  by v1.2.1 or before.

·         The software cannot open files from ViiA™ 7, StepOne Plus®, 7900 HT Fast, 7500 Fast, or 7500 Instruments.

·         The software can open setup files created by 7900 HT Fast SDS Software v2.3.

·         The software cannot communicate/network with other non-QuantStudio™ 12K Flex instruments.

 

 

KNOWN ISSUES

Security, Auditing and E-signature

·         In a secure environment with auditing enabled, if you run an experiment created from a template in automation mode, saving the experiment is not audited. If you need to have the save appear in the audit records, create experiment files using the Batch Experiment Setup feature before running the experiments in automation mode.  This will list the experiment in the audit record.

·         Disabling audit and e-signature setting in the secured environment does not hide/disable the Audit experiment menu.

·         In a secure environment with auditing enabled, Audit Report does not include Comments field.

·         When double-clicking on an .EDS file from the operating system to launch the application, the system will not open the experiment if the .EDS file does not contain audit records when Auditing is enabled and the Allow Open of non-audited EDS files setting is also enabled.  Launch the application first and open the experiment from within the application.

·         Enabling or disabling e-Signature in the secured environment requires the application to be restarted to take effect.

 

 Calibration

 

Analysis Setting

 

Viewing Analysis Results

·         There are some cases where the completed run analysis results have loss cycle data in the Amplification Plot. When this happens, we recommend that you reanalyze the run data by selecting “Analysis from Raw Data” from “Analysis” menu.

 

 

 *****Known Issues for OpenArray® Blocks *****

 

Performance

·         System can take up to 3 minutes to perform each of the following operations:

o   Opening an OpenArray® real-time genotyping experiment

o   Analyzing an OpenArray® block experiment

o   Fails to detect OpenArray® plate center (no warning message is displayed)

·         Modifying analysis settings and re-analyzing with many wells selected (e.g. entire experiment) will give the appearance that the application has frozen for up to 6 minutes.  Therefore, unselect wells before modifying analysis settings.  Use bookmark feature to remember selected wells.

 

Import / Export / Report

·         The date format in report is always shown as yyyy-MM-dd.

·         The system is unable to export automatically upon completion of an OpenArray® block run.  Therefore, open experiment file after the run has completed and export manually.

·         Legend not shown fully in report and in an exported image file.

·         Export with Filter Bookmark Data selected as ‘Yes’ incorrectly exports the previewed data from the Amplification tab when 20 or less wells are bookmarked.  Therefore, bookmark more than 20 wells to export correct data.

·         Export with Filter Bookmark Data selected as ‘Yes’ incorrectly exports the wrong data from both the Raw Data and Multicomponent tabs.  Therefore, export with Filter Bookmark Data selected as ‘No’ to export correct data.

·         Instrument Details tab is not included in the Export.

·         Only general calibration status is included in export.  Calibration status for each calibration type is not included.

·         Auto-exported .xlsx files cannot be opened by the application.  The work around is to export manually after the run.

 

Plot

·         EMF format is not supported in this release of the software.

·         Data points in CT vs. Well Plot of OpenArray® block experiment cannot be filtered based on user-selection.

·         Disabling Auto-adjust Range from Plot Properties of Allelic Discrimination Plot does not work.  Therefore, do not disable Auto-adjust Range setting.

·         Create Slide operation for HRM experiment does not include pre- and post-melt range on the plot.

 

Settings

·         Preferences settings of Sample Tracker software only take effect after software is re-launched.

 

Installation

·         Sample Tracker software will not automatically install when QuantStudio™ 12K Flex software is not installed in C:\ drive.

 

Instrument Run

·         Due to high utilization in the instrument CPU:

o   Periodically, instrument screen is unresponsive.  Countdown timer will function properly.  Instrument status light may flash amber.  Run continues appropriately and can be monitored remotely from a software client computer.  Experiment files can be downloaded from the software.

o   Periodically, the instrument screen freezes.  Countdown clock will stop.  Run continues appropriately and can be monitored remotely from a software client computer.  Experiment files can be downloaded from the software.

·         Instrument may not boot up correctly if power is interrupted during firmware upgrade.  Re-install firmware to ensure proper operation of instrument.

·         Instrument display of real-time amp plot during a run may differ slightly from the software display due to additional processing applied in the software.

·         When performing an instrument run from a Post-Read Genotyping template (.EDT), system will ignore the Pre-Read stage and Amplification stage settings in Preferences for OpenArray® blocks.

 

Chip Identification

·         Chip detection at the beginning of a run can take up to 3 minutes, although typical processing period is approximately 40 seconds.

·         If less than four OpenArrays® plates are loaded for an instrument run, the Software will display a progress bar dialog indefinitely.  Click the ‘X’ to close the dialog, or include old OpenArray® plates in the carrier to fill the empty positions.

 

RNaseP Calibration

·         User is unable to omit wells from the Plate Layout view.  Select wells and navigate to Results table to omit wells.

·         Omitting wells and then clicking ‘Finish’ without reanalyzing will display empty values for Amp Score and CRT.  To view the actual values, transfer the experiment manually from the Instrument Console, then open and reanalyze the experiment.

·         The system intermittently will not refresh the amplification plot upon completion of a run.  To view the plot, close and re-open the RNase P calibration screen from the Instrument Console.

 

Connecting to Multiple Instruments

·         It is advised to monitor only one OpenArray® block run from a single computer due to the large volume of data being processed.

 

QC Summary

·         The QC Summary screen for an OpenArray® plate experiment is extremely slow to access.  For quicker results, refer to the QC Summary screen in either ExpressionSuite Software v1.0, TaqMan® Genotyper Software v1.2 or DigitalSuite™ Software v1.0.

 

Experiment Status

·         The status for a completed action for an experiment remains in the status bar when navigating away from that experiment.

 

Automation/Batch Experiment Utility

·         Slow performance is experienced after creating a large number of experiments using the batch utility. To work around this, turn off auditing when creating new experiments from the batch utility, or re-start the Software after executing the batch utility once or twice.

·         The system may run out of memory, particularly when Security is enabled, after repeated execution or when large numbers of experiments are used for Batch Experiment Setup operation.  To avoid this, limit the number of experiments to 50 and restart application after use of this operation.

 

Opening Experiment Files

·         The system may run out of memory after repeated opening and closing of experiment files.  Close and re-open the software application to continue using the software.

·         The system will not open an OpenArray® genotyping experiment with a pre-read stage when transferred manually after the run from the Instrument Console.  The work around is to transfer the experiment manually via USB stick.

 

 

*****Known Issues for non-OpenArray® Blocks *****

 

Instrument Run

·         If a run is terminated during an infinite hold, system incorrectly labels experiment with status of “Run Terminated,” rather than “Run Completed.”  System is able to open and analyze file correctly.

·         For some experiment types created from Create From Template operation, real-time amplification plot will not display during the run.

 

Calibration

·         In Background calibration, wells that failed QC are not highlighted to the user in the plate layout.  Navigate to Well Table to view calibration results.

·         When the instrument is disconnected from the software just before a calibration run, there may be multiple error messages displayed. When this happens, check the Instrument Console to ensure the instrument is in the “Ready” state before trying again.

·         Calibration runs may return a QC warning message in the QC tab that describes a failure condition.  Disregard the warning message.

·         ROI calibration run does not fail when using a SYBR® dye plate.  To avoid this scenario, do not use SYBR® dye plate for ROI calibration run.

 

Connecting to the Instrument

·         The status of the instrument remains as "Not Connected" in the Instrument Console after the IP address of the instrument has changed. Restart the software after changing the IP address of an instrument.

 

Notifications

·         Calibration reminder settings cannot be tested from the calibration reminder screen. Therefore, to test email notification and SMTP settings, create a new experiment and test the notification settings from the experiment run screen.

·         Email notification operation incorrectly does not function with SMTP encryption enabled.  Uncheck the Encryption Required setting in Preferences for SMTP Settings to allow notifications.

 

Genotyping Experiments

·         The initial view of the allelic discrimination plot does not show the different colors for each of the genotypes. Click on the empty region of the plot to see the genotype colors.

·         For 96-well 0.2 mL and 384-well block types only, when creating a new Genotyping experiment without using a template, the Reaction Volume per Well is incorrectly populated by default as 50 µL and 20 µL, rather than 20 µL and 5 µL, respectively.  The work around for these two block types is to manually edit the Reaction Volume per Well from the Run Method screen prior to saving the experiment.

 

Experiments

·         If you open more than 10 experiments at the same time, the bottom right arrow to move the tab becomes invisible. Close some experiments to assess the hidden ones.

 

Import and Export

·         Importing an .AIF file into a genotyping experiment does not import the context sequences. Therefore, import an .AIX file into a genotyping experiment to import the context sequences correctly.

·         Importing plate setup file with Quencher field set to ‘None’ or ‘NONE’ may corrupt EDS file.  Once saved, user may not be able to re-open .EDS file. To avoid this, use ‘NFQ-MGB’ or any valid dye name as the Quencher.  Recommended safeguard is to make a backup copy the original .EDS file prior to importing plate setup.  Contact technical support if your data becomes corrupted. 

·         To export raw data for a Standard Curve experiment that includes a melt curve, user must click Load Export Set before exporting the data.

 

Viewing Analysis Results

·         There are some cases where analysis results are mislabeled with ‘CT’, even though the Relative-Threshold algorithm was used. To be certain which algorithm was used to calculate the results, open the Analysis Settings dialog box and review the analysis settings.  While the CT labels are incorrectly displayed, the results are correct for the Relative-Threshold algorithm.

·         Displayed results of CT Threshold and Amp Score incorrectly shows three decimal places when Decimal Places to Show setting in Preferences for Non-Array is other than 3.

·         For ddCT experiments only, displayed results incorrectly excludes Amp Score, dCT, Baseline Start, Baseline End and Number of Replicates, and incorrectly includes CT Mean, CT SD and dCT Mean when RQ Settings for Efficiency is set to 100%; and incorrectly excludes CT Mean and CT SD when Efficiency is other than 100%.

·         Upon completion on an instrument run, system will not automatically transition to Analysis screen for Presence/Absence, Genotyping, HRM and Melt Curve experiments.

 

Presence/Absence

·         False presence calls. Due to the enhanced uniformity of the instrument, the threshold for the Presence/Absence call may be too close to the NTC wells. Increase the confidence level from 99% to move the threshold farther away from the NTC wells.

 

Print Report

·         The Experiment Results Report for a genotyping experiment always displays the column headers ‘Allele 1 CT’ and ‘Allele 2 CT’ using the text ‘CT’, even if the Relative-Threshold algorithm was used. To determine from the report which algorithm was used to calculate the results, refer to the Quantification Cycling Method on page one of the report.  While the CT labels are incorrectly displayed, the results are correct for the Relative-Threshold algorithm.

 

 High Resolution Melt Experiments

·         For a High Resolution Melt experiment, manual calls cannot be made in the Variant Call column in the Well Table. To make a manual call, double-click a well in the Plate Layout tab and select the control from the Control drop-down list.

·         For a High Resolution Melt experiment, the Derivative Melt Curve plot in the Run area is empty. To view the derivative melt curve, click Analysis, then HRM Plots.

·         Exporting with more than 65,536 rows is not supported by the Excelâ 2003 format. To export such data use either the Text format or the Excelâ 2007 format.

·         Recommended hard drive space available when performing an instrument run for a High Resolution Melt experiment is 50MB.

 

 

SYSTEM REQUIREMENTS

The computer hardware and operating system requirements for the QuantStudio™ 12K Flex Software are:

·         Windowsâ 7, 32-bit with Service Pack

·         Pentium 4 or compatible, with minimum 2GB of RAM and 20 GB of hard drive capacity

·         Minimum monitor resolution of 1280x1024

·         One open Ethernet port for connecting to the instrument directly

·         Internet Explorerâ 6.0 or higher (for online assay browsing and ordering)

·         Microsoftâ PowerPointâ software (for direct export into PowerPoint slides)

·         Microsoftâ Excelâ software

  

 

INSTALL SOFTWARE

IMPORTANT - To help prevent data loss, it is strongly advised that all user data is backed up before upgrading the software.

·         Log on to the Windows system with Administrator privileges.

·         Obtain the software installation package.

·         Place the disk in the CD drive and allow the installer to launch automatically.  If the installer does not launch automatically, double click the setup.exe file on the CD to start the installation.

·         Agree to the End User License Agreement (EULA) when prompted and complete the installation.

·         The QuantStudio™ 12K Flex Software v1.2.2 requires a license key for activation.  Follow the instructions in the installation wizard to enter your license key.

·         The wizard directs you to a QuantStudio™ 12K Flex Software license activation URL (https://licensing.appliedbiosystems.com/activation/QuantStudio12KFlex) to receive your license file.

·         After you have your license file, during the installation process, navigate to location of the license file to activate the software.

·         Your license file, while permanent, must be renewed every two years.

·         The software will be installed, by default, to “C:\Applied Biosystems\QuantStudio 12K Flex” (referred to as the home directory of the QuantStudio 12K Flex Software).

·         A program group, “Applied Biosystems/ QuantStudio 12K Flex Software” will be created during installation. In addition, a short cut to the application launcher, “QuantStudio 12K Flex Software ”, will be installed on the computer desktop.

·         When a previous version of the software is detected on the system, the installer will perform an upgrade. The user will be prompted to proceed with the upgrade or to cancel the operation.

 

 

INSTALLATION OF NEW MODULES

·         If you have purchased additional modules such as High Resolution Melt Module or Security and Auditing for QuantStudio™ 12K Flex Software, a license key is required to activate the modules.

·         Select Tools > License Central from QuantStudio™ 12K Flex Software.

·         Follow the instructions in the wizard to enter your license key. The wizard directs you to a QuantStudio™ 12K Flex Software license activation URL (https://licensing.appliedbiosystems.com/activation/QuantStudio12KFlex) to receive your license file. 

·         After you have your license file, during the installation process, navigate to location of the license file to activate your module. 

·         Restart the QuantStudio™ 12K Flex Software after the license has been successfully applied.

 

 

UNINSTALL SOFTWARE

IMPORTANT: To help prevent data loss, it is strongly advised that all user data is backed up before uninstalling the software.

·         From Windows “Start” button, find the “Applied Biosystems/QuantStudio 12K Flex Software” program group.

·         Click on “QuantStudio 12K Flex Maintenance”.

·         In the wizard, follow the instructions and complete the uninstall operation.

·         Optionally, rename or delete the home directory of the QuantStudio 12K Flex application. This ensures a clean environment for the next installation.

  

 

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