GDS Basic

Assurance GDS
Assurance GDS utilizes the latest technology developments to
achieve unprecedented time to results and Accuracy
 IMS-based sample preparation
 Highly specific primers and probes
 Innovative rotary instrument platform
Advances in multiple areas
Sample
Preparation
Reagent
System
Instrument
Platform
Accuracy + Speed + Ease of Use
Sample Preparation
Immunomagnetic Separation (IMS)
What is IMS:
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“Immunomagnetic Separation”
Antibody specific capture and separation of organisms
Capture – antibody coated paramagnetic particles
Separation – magnet collects and isolates particles with bound
organisms
Immunomagnetic Separation (IMS)
Improves purity of sample:
 Reduces competitive microflora
 Incremental increase in specificity
 Improves isolation of target organism on agar plates
 Improves selectivity of enrichments
 Removes potential PCR inhibitors
 Food matrix, sanitizers, culture media, etc
Increases quantity of DNA for Analysis
 Concentrates target organisms
 Provides a 1-2 log increase in cell density post-enrichment
 Analysis of larger test portions (1,000 µL vs. 5 µL)
Assurance GDS
Sample Preparation
IMS Made Easy:
1.
Add 1mL of enriched sample to IMS
particles
2.
Mix 5-10 minutes
3.
Transfer particles through wash solution
to resuspension buffer with PickPen
4.
Transfer to amplification tubes
5.
Amplification and detection with
Assurance GDS Rotor-Gene (75 minutes)
Reagent System
Detection
Probe-based
detection
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Concurrent amplification and detection
Sequence specific
Additional specificity
Not affected by extraneous DNA
Discrete signal for target and IC
How the MGB Eclipse Probe Works
hv
= Fluorophore
Probe in solution
 Arranged in a random coil
 Fluorescent marker quenched
= Quencher
Probe bound to target
 Probe becomes linear
 Fluorescent marker exposed
= MGB
MGB
Direct correlation between the amount of fluorescence and the amount of target
Assurance GDS
Reagent System
Benefits Assurance GDS Probe

MGB – improved specificity
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Allows for shorter target sequences
Prevents mismatches
Better discrimination of target
No melt-curves
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Saves > 1 hour of instrument time
No subjective interpretation
Assurance GDS
Reagent System
Benefits Assurance GDS Probe

Target specific detection
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True internal control
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distinct primer and probe set
Separate signals
Direct detection of target
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1:1 relation of probe to target
Instrument Platform
Traditional
Thermocyclers
Traditional Format
 Peltier block based heating / cooling
 Single channel detection
Limitations
 Slower cycling – increased dwell
times between temp. changes
 Inconsistent temperature cycling
between wells
 Dependent upon non-specific dye
and melt-curve analysis
Assurance GDS
Instrument Platform
Assurance GDS Rotor-GeneTM
 Samples arranged in a rotary format for direct
heating / cooling
 Centrifugal motion with constant air exchange
 Ensures uniform temperature among all
reactions
 Eliminates lengthy dwell times required by
block based systems
Assurance GDS
Instrument Platform
 Rotor-Gene controlled
PCR reactions are highly
consistent
 Produces accurate &
reliable test results
# of cycles
Improved Accuracy
 Generates confidence
Test #
Assurance GDS
Instrument Platform
Faster Results
PCR Step
Rotary
Block
Time savings
Starting denaturation time
3 min
10 min
7 min
Typical cycle time
<2 min
4 min
--
Total cycle time ( 42 cycles)
70 mins
170 min
100 min
0
60 min
60 min
75 min
240 min
>2.5 hours!
Melt Curve
TOTAL
Pathogen Detection in Practice
Assurance GDS family of assays:
• E. coli O157:H7
• Shiga Toxin Genes
• Salmonella
• Listeria spp.
• Listeria monocytogenes
• Enterobacter sakazaki (coming soon)
Assurance GDS for E. coli O157:H7
AOAC Official Method 2005.04
Protocol:
mEHECTM
(6 ½ - 18 hrs)
PickPen IMS sample prep
(20-30 minutes)
Amplification & Detection
(75 minutes)
Total Time to Results – 8 hrs
Assurance GDS for Shiga Toxin Genes
AOAC Official Method 2005.05
Protocol:
mEHECTM
(8 - 18 hrs)
PickPen IMS sample prep
(20-30 minutes)
Amplification & Detection
(75 minutes)
Total Time to Results – 10 hrs
Assurance GDS for Salmonella
AOAC PTM (050602)
Protocol:
Buffered Peptone Water
(18 - 24 hrs)
High Microbial Foods
Low Microbial Foods
PickPen IMS sample prep
(20-30 minutes)
PickPen IMS sample to
BHI (2-4 hrs)
Amplification & Detection
(75 minutes)
Total Time to Results: 20-24 hrs
Assurance GDS for L. monocytogenes
AOAC PTM 070702
Protocol:
Environmental Samples / Select Foods
BLEB + LiCl + PALCAM
(18-24 hrs) + (4-8hrs)
Other Foods
Demi-Fraser Broth
(30-48 hrs)
PickPen IMS sample prep
(20-30 minutes)
Amplification & Detection
(75 minutes)
Total Time to Results: 24 hrs (Environmental / Select Foods)
32 hrs (Other foods)
Assurance GDS for Listeria spp.
AOAC PTM 070701
Protocol:
Environmental Samples
BLEB + LiCl + PALCAM
(18-24 hrs) + (4-8hrs)
Food
BLEB + LiCl + PALCAM
or Demi-Fraser Broth
(30-48 hrs)
PickPen IMS sample prep
(20-30 minutes)
Amplification & Detection
(75 minutes)
Total Time to Results: 24 hrs (environmental)
32 hrs (food)
Assurance GDS for Enterobacter sakazakii
Protocol:
Cronobacter Broth
(24-28 hrs)
Sample prep (20-30 minutes)
Amplification & Detection
(75 minutes)
Total Time to Results: 26 hrs

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