next generation panel oriented diagnostics the filmarray pathogen identification system idaho...
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NEXT GENERATION PANEL ORIENTED DIAGNOSTICS THE FILMARRAY PATHOGEN IDENTIFICATION SYSTEM
Idaho Technology
The FilmArrayThe FilmArray
The FilmArray is…The FilmArray is…
User-Friendly Multiplex PCRUser-Friendly Multiplex PCR
Automated ProtocolAutomated Protocol
Integrated Sample PrepIntegrated Sample Prep
Multiplex PCRMultiplex PCR
Automated Results AnalysisAutomated Results Analysis
SpeedSpeed
Load Pouch
Run Set-upRun Set-up
Inject pouch Hydration Solution
Add sample to Sample
Buffer
Run Set-upRun Set-up
Inject sample
Enter pouch and sample ID, user info, then
press go!
Load pouch in FilmArray
Automated ProtocolAutomated Protocol
Automated ProtocolAutomated Protocol
5 min of hands on time
Start the run &Start the run &
walk awaywalk away
Integrated Sample PrepIntegrated Sample Prep
Multiplex PCR
Simultaneous Simultaneous detection of detection of dozensdozens of targets of targets from one from one patient samplepatient sample
Automated Results AnalysisAutomated Results Analysis
Positive or negative Positive or negative call for each targetcall for each target
Automated Results AnalysisAutomated Results Analysis
Eliminate SubjectivityEliminate Subjectivity
Speed…. It’s really fast.Speed…. It’s really fast.
Sample prep
Results
Multiplex PCR
Analysis
RT-PCR 1 hour1 hour
The FilmArray
How does it work?
The FilmArray PouchThe FilmArray Pouch
Reagent Reagent StorageStorage
Chemical Chemical Circuit Circuit BoardBoard
Sample Sample extraction & extraction & purificationpurification
11stst stage stage multiplex multiplex
PCRPCR
22ndnd stage stage PCRPCR
The FilmArray – The FilmArray – How Does it Work?How Does it Work?
FilmArray Animation
Automated Protocol
Pneumatic Bladders
Bladders inflate over blisters to push liquidAir Channels
Pistons close channels
The FilmArray: Video analysisThe FilmArray: Video analysis
Integrated Sample PrepIntegrated Sample Prep
+ + =
Nested MultiPlex PCRNested MultiPlex PCR
Automated Results AnalysisAutomated Results Analysis
102 individual 2nd stage PCR wells
Each well contains one reaction
Fluorescent data is generated for each well
Automated Results AnalysisAutomated Results Analysis
All targets tested in triplicate
Automated Results AnalysisAutomated Results Analysis
For some organisms we target multiple genes – each tested in triplicate
Software generates an automated call for each organism
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The FilmArray Respiratory Panel
ViralAdenovirus Bocavirus Coronavirus 229E Coronavirus HKU1 Coronavirus OC43 Coronavirus NL63 Entero VirusInfluenza A Influenza A H1Influenza A H1 2009Influenza A H3
Bacterial Bordetella pertussis Chlamydophila pneumoniae Mycoplasma pneumoniae
Influenza B Metapneumovirus Parainfluenza 1 Parainfluenza 2 Parainfluenza 3 Parainfluenza 4 Respiratory Syncytial Virus Rhinovirus
Nested Multiplex PCR
Blue Graph - 5000 x LOD* C. burnetti
Red Graph - 0.1x LOD* B. anthracis (100 ul of a 100 CFU/ml BA sample)
Comparison of Multiplex PCR
*LOD = 1000 cfu/ml
Robust Multiplex PCR
Triple Infection
Bocavirus – black
PIV2 – blue
Human Rhinovirus - green
Bocavirus – black
PIV2 – blue
Human Rhinovirus - green
Simultaneous Detection of 12 OrganismsSimultaneous Detection of 12 Organisms
Flu A H1Flu A H3Flu BRSVPara Flu 1Para Flu 2
Para Flu 3AdenoHuman Meta PneumoRhino Boca S. pneumoniae
System Controls
Controls currently in the pouch:RNA Process mRNA from freeze-dried S. pombe
Pouch pass/fail is set on this control
1st Stage PCR synthetic oligos in PCR1
2nd Stage PCR synthetic oligos in PCR2
Useful for understanding failure modes
Human mRNA assay for human mRNA in sample
We are collecting data
Internal pouch Controls
Location of the controls
PCR2 control (spotted with PCR2 primers)
PCR1 controlFreeze-dried with Primer Pellet
Yeast controlFreeze dried into sample well
Comparative Data
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The FilmArray – Data
FilmArray vs DFA in 387 Pediatric NPAs
Pediatric NPA samples were tested by DFA (blue) and by FilmArray (maroon).
82% of 387 pediatric NPA samples tested 82% of 387 pediatric NPA samples tested positive by FilmArray analysispositive by FilmArray analysis
32% tested positive for 2 or more 32% tested positive for 2 or more pathogens by FilmArray analysispathogens by FilmArray analysis
Future Developments
Product Development (Beyond RP)
Sepsis/Meningitis Panel
Enteric Pathogens Panel
STD Panel
Biothreat Panel
Questions