GenMark Diagnostics...Training Presentation GNMK-IMC-6011-D . Training Agenda Thrombophilia Risk...
Transcript of GenMark Diagnostics...Training Presentation GNMK-IMC-6011-D . Training Agenda Thrombophilia Risk...
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GenMark Diagnostics
eSensor® Thrombophilia Risk Test (IVD)
GNMK-IMC-6011-D
Training Presentation
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Training Agenda Thrombophilia Risk Test
• Test Overview
• Kit Components and Handling
• Assay Setup
• Reporting
• Quality Control
• Ordering and Contact Information
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Test Overview
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Thrombophilia Risk Test (IVD) Intended Use
The eSensor® Thrombophilia Risk Test
is an in vitro diagnostic for the detection
and genotyping of Factor II
(Prothrombin), Factor V (FV Leiden) and
MTHFR (methylenetetrahydrofolate
reductase gene) C677T and A1298C
mutations in patients with suspected
thrombophilia.
*The complete intended use statement may be viewed in the Package Insert, which can be
found in the Customer Training Binder or on the GenMark Customer Resource Center.
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Thrombophilia Risk Test Get the complete picture of inherited thrombophilia risk
• 100% accuracy and
reproducibility in clinical trials
• Single tube multiplex test for all
four thrombophilia-related
polymorphisms
• Test results in 4 hours including
extraction − ~40 minutes of hands-on time*
• Objective results automatically
generated by the XT-8 system
Gene Polymorphism
FV Leiden 1691G>A
FII Prothrombin 20210G>A
MTHFR 677C>T
1298A>C
* Based on actual workflow studies conducted by GenMark scientists at customer sites. Times used are averages across multiple
sites for calculations.
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Kit Components and Handling
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Thrombophilia Risk Test Components Each kit contains components and reagents for 48 tests
eSensor Thrombophilia Risk Test Package Insert can be downloaded from GenMark’s Customer Resource
Center at http://genmarkdx.com/customer_resources.php.
Kit Pack/Boxes Component Packaging & Quantity Storage
Cartridge Pack Thrombophilia Risk Test
Cartridge Pouch
6 foil bags with 8 cartridges in
each bag 10 – 25 °C
PCR Box
TRT PCR Mix 2 vials containing 1100 µL each -20 °C
(Pre-PCR Location) Taq Polymerase 1 vial containing 60 µL
Genotyping Box
Exonuclease 2 vials containing 145 µL each
-20 °C
(Post-PCR Location)
TRT Signal Buffer 2 vials containing 2500 µL each
Buffer-1 2 vials containing 350 µL
Buffer-2 2 vials containing 700 µL
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Lot Number and Storage Information Package location
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Reagent Storage, Handling, and Stability
• Do not pool reagents from different lots.
• Immediately after use, close all vials to prevent spillage or
contamination.
• Store all vials in an upright position.
• Reagents may be frozen and thawed up to 5 times.
• Cartridges must be used within 30 days of opening the pouch.
• Do not use a kit after its expiration date.
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Assay Set Up
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Thrombophilia Risk Test Workflow Overview
Total assay time of 4 hours with ~40 minutes of hands-on time
* Based on actual workflow studies conducted by GenMark scientists at customer sites. Times used are averages across multiple
sites for calculations.
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Load PCR
reactions into
thermal cycler
Add 5 µL of
exonuclease
to each sample
PCR
Dispense sample
and hybridization
solution into
cartridges
Load cartridges onto
the eSensor® XT-8
system(s)
Generate
reports
Thrombophilia Risk Test Workflow Overview
Capable of high-
throughput testing
using a 96-well
plate format
OPTIONAL
HOLD POINT
OPTIONAL
HOLD POINT
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dsDNA
Amplicon
40 µL
EXONUCLEASE
DIGESTION
TRT Signal Buffer
70 µL
Buffer 1
10 µL
Buffer 2
20 µL
Hybridization
Solution
100 µL
ssDNA
Amplicon
140 µL
GENOTYPING
SETUP
Genomic DNA
5 µL
PCR Reaction Mix
35 µL
PCR SETUP PCR Master Mix
30 µL
PCR Mix
29 µL Taq Polymerase
1 µL
125 µL
eSensor® XT-8
Cartridge
HYBRIDIZATION / DETECTION
EXONUCLEASE SETUP Exonuclease
5 µL
PCR
100 µL
Thrombophilia Risk Test Detailed Workflow
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Exonuclease Technique
X
Exo addition Post-vortexing Post-centrifuging
NOTE: Dye was added to improve visualization of exonuclease and is not added to the
exonuclease supplied with the kit
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Proper Cartridge Insertion Technique
X
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Entering Lot Information Scan the barcode into the XT-8 system
• Frozen reagents are shipped in a
ziplock bag labeled “eSensor
Thrombophilia Risk Test Reagents”
• 2D barcode on label is for GenMark
Internal Use Only and can’t be used
for scanning lot information into the
XT-8
• Use the barcode found on the side of
the Detection Reagents box to enter
lot information
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Procedural Notes • General
– Dedicated pipettes, tips and equipment are recommended for pre and post-amplification activities
– Unidirectional laboratory workflow is recommended
• Reagents – Frozen reagents and samples are thawed at 37 °C for 10-15
minutes or at room temperature
– Mix using a vortexer at maximum speed for 3 - 5 seconds
– Consolidate liquids by brief centrifugation (5-10 seconds) in a centrifuge or mini-centrifuge
– Do not thaw or vortex Taq Polymerase or exonuclease enzymes!
• Samples – Whole blood colleted with EDTA as anticoagulant
– Frozen whole blood is not recommended for use with this product
– Genomic DNA Concentration:
2 ng/µL (10 ng DNA/PCR) - 100 ng/µL (500 ng DNA/PCR)
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Reporting
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Sample Report
• Single-page report
• Results Section lists determination made for each of the four polymorphisms
• Customized reporting options
• Reflex report option – No additional testing required
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30
• Full Panel (FV, FII, MTHFR 677 & 1298)
• FII
• FV
• FII/FV
• MTHFR 677 & 1298
5 Report Types
Reflex Report Option- No need to run individual tests for each desired result
Customized Report Options
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Reporting Viewing Reports
• Reports can be filtered according
to different parameters:
− Accession Number
− Cartridge Lot
− Date (see screen shot)
− Protocol
− User
− Batch
− Printed Reports
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Reporting Viewing reports by accession number
• Select the desired
accession number(s) - Multiple accession numbers
can be selected for a
particular report type
• After accession number(s)
is selected, chose the
Report Type by touching
the Report Type button
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Reporting Selecting the Report Type
• Users can select from 5 report
types on the Report Type list: − Full Panel:
(FV, FII, MTHFR 677 & 1298)
− FII
− FV
− FII/FV
− MTHFR 677 & 1298
• Once Report Type is chosen,
use the touch screen buttons
to view, print, or export the
report
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Reporting Reflex to MTHFR 677_1298
• To reflex to MTHFR results,
select the corresponding
accession number(s)
- Multiple accession numbers
can be selected
• After accession number(s)
are chosen, select the
“FV_FII_677_1298” Report
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Reporting Selecting the _677_1298 Report Type
• To view only MTHFR
results, select the
“_677_1298” report type
• Use the touch screen
buttons to view, print, or
export the report
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Quality Control
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Quality Control
• Training Samples – QC samples are provided for use during training, and can be used
as control material during validation
• QC Vendors
– Coriell Institute
• www.Coriell.org
• Phone: 1-856-966-7377
– Maine Molecular Quality Control Institute (MMQCI)
• www.MMQCI.com
• Phone: 1-207-885-1072
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• On-Board Positive Control
– Hybridization Failure
– Detection Failure
• On-Board Negative Control
– Assay System Failure
• Genotyping Metric – Monitor and trend performance of test
– Shows distance of the genotyping score from the nearest relevant boundary (indeterminate zone)
– Acceptable range 0.9 – 1.0
Quality Controls On-Board
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Quality Controls No Template Control
• PCR Blank (DCM Report)
– Method to monitor for amplicon contamination during
setup of the reaction
– Use Nuclease-free water (Blank)
– Summary Result:
• Metric Value: 0.0 – 1.0
VALUE FOLLOW UP ACTION
0.0 No Action
0.1 – 0.8 Monitor for trend of increasing value
0.9 – 1.0 Decontaminate work areas and repeat entire run
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Ordering and Contact Information
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Thrombophilia Risk Test Ordering Information
• Item # MT004018
• Kit Configuration 48 tests per kit
• Regulatory Status IVD
• Customer Service
Telephone 1-800-eSensor (373-6767), Option 1
Email [email protected]
Fax Number 1-866-831-2001
• Technical Support
Telephone 1-800-eSensor (373-6767), Option 2
Email [email protected]