melissa may netbio - vice president – strategic planning date: 22/10/2013
TRANSCRIPT
Melissa May
NetBio - Vice President – Strategic Planning
Date: 22/10/2013
Heading
Fully automated, Field forward Rapid DNA Typing for Military, Intelligence, and Law Enforcement Applications
090413
Requirements: ANDE Rapid DNA Analysis System (I)
In 2009 NetBio was awarded the ANDE contract, a competitive R&D program funded by a consortium of 5 Federal agencies to develop a Rapid DNA Analysis System based on:
• Ease-of-use. The Operator has no technical training.– No manual processing– No reagent loading—reagents should be incorporated into the single-
use consumable– No instrument calibration or assembly– No data interpretation—automated allele calling and searching
• Reagent Stability/Out-of-the-box functionality. The System will be placed in a field forward environment. – No temperature, humidity, or particulate control – No refrigeration– No tools or technicians to install/align system – Generator power
• Ruggedization. The system must be hardened for transport to and operation in deployed environments.
• Rapid. DNA analysis must provide results within a time period that allows decisions to be made BEFORE releasing a person of interest.
• Data security. All aspects of system operation and data generation must be secure.– User and sample tracking– Encryption of all generated data and database
• Flexibility. The system must provide a customizable foundation for field forward forensic DNA analysis.– Touch sample analysis– Kinship analysis– Multi-assay capability
Requirements: ANDE Rapid DNA Analysis System (II)
ANDE System Components and Process Flow
Collection Swab
Fully AutomatedInstrument
BioChipSet™ Cassette
AutomatedData Analysis
ANDE System Specifications
• Sample Types: Fresh and dried buccal and blood swabs, purified DNA.
• Number of Samples Per Run: 5 plus allelic ladder.
• Analysis Time: DNA profile in 84 minutes.
• Operator Responsibility: Insert samples and initiate run. No other manipulations are required.
• Reagent Stability: Reagents are stable for 6 months at room temperature, no refrigeration required.
• Data Analysis: Fully automatic, integrated expert system software. No analyst review required.
• Sample Tracking: Integrated RFID scanner on board the instrument allows tracking of RFID enables sample swabs.
• System Maintenance: Field serviceable, annual maintenance.
• Data Security: Password protected access and data encryption.
Operating the ANDE System: Collect Samples
Enter Sample ID
Scan a Sample with the Integrated RFID Reader
Insert Sample and Press the Done Button
Load the Sample into the BioChipSet
Press the Done Button
Insert the BioChipSet Cassette
Initiating the Run
DNA Processing Complete
Processing Complete
Female DNA Profile
STR Analysis, How Does It Work?
Known: Reference Sample:• Convicted Offenders• Arrestees• Suspects• Paternity
Unknown: Evidence Sample:• Blood, Body Fluids• Cigarette Butt• Handled Objects (cup, pop can, water bottle, etc)• Murder Weapon
DNA Purification
DNA Quantitation
STR Amplification
CE Separation & Detection
DNA Profile Generation
DNA Purification
DNA Quantitation
Database Search/Match
STR Amplification: What Are Short Tandem Repeats (STRs)?
• Human DNA is 99% identical from person to person. However, some regions of the genome vary, and one type of variable region is the Short Tandem Repeat (STR).
• Short Tandem Repeats (STR) are repetitive sequences:– Tetranucleotide: AATG AATG AATG AATG
7 repeats (maternal)
8 repeats (paternal)
AATG
~
~
~
~
STR Analysis, How Does It Work?
Known: Reference Sample:• Convicted Offenders• Arrestees• Suspects• Paternity
Unknown: Evidence Sample:• Blood, Body Fluids• Cigarette Butt• Cup• Murder Weapon
DNA Purification
DNA Quantitation
STR Amplification
CE Separation & Detection
DNA Profile Generation
DNA Purification
DNA Quantitation
Database Search/Match
STR Amplification
Female DNA Profile
DNA Profiling is Extraordinarily Accurate
The probability of a random match for this STR profile in the U.S. population is less than 1 in 837 trillion
J. Bulter, http://www.cstl.nist.gov/biotech/strbase/fbicore.htm
Locus Allele Value Allele Value 1 in Combined
D5S818 11 0.2533 13 0.2152 9.17 9
VWA 17 0.2815 17 8.87 81
FGA 25 0.1854 26 0.2185 12.35 1005
D8S1179 11 0.1854 14 0.1656 16.29 16,364
D21S11 28 0.1589 31 0.2782 11.31 185,073
D18S51 13 0.1374 17 0.1391 26.18 4,845,217
D3S1358 15 0.3841 16 0.1407 9.25 44,818,259
D13S317 10 0.3394 14 0.0480 30.69 1.38 x 109
D7S820 10 0.1772 10 31.85 4.38 x 1010
D16S539 9 0.1126 13 0.3212 13.80 6.05 x 1011
THO1 6 0.2318 9 0.2814 18.62 1.13 x 1013
TPOX 8 0.5348 8 3.50 3.94 x 1013
CSF1PO 11 0.2169 12 0.3212 21.28 8.37 x 1014
Traditional DNA Forensic Laboratory Infrastructure
Sample Prep Lab
Amplification Lab
Separation & Detection Lab
Forensic AnalysisU.S. Crime Lab $210 M, 351,000 sq ftDNA Unit ~$56 M, 70,000 sq ft, annual operational budget >$2 M
• Laboratory Infrastructure• Capital Equipment• Lab Supplies and Reagents • Computers and software• Laboratory Technicians• Forensic Analysts
Five Lab Processes in One Portable Device
Rapid DNA Analysis for Human Identification is the automated generation of STR profiles (DNA fingerprints) by non-technical operators outside the laboratory.
Human AnalystProfile Match
Hours - Weeks
ANDE: NetBio’s Rapid DNA Instrument
• Instrument Weight: 50 kg (two person lift)• Instrument Size: 68 x 42 x 59 cm• Ruggedized to MIL STD 810F for shock / vibration• No calibration or realignment required after transport
RFID Reader
Touchscreen Monitor
BioChipSet Access Door
Folding Carry Handle
GPS Window
Field Forward Testing to MIL STD 810F
The ANDE System has under gone extensive ruggedization and field testing
• Shock
• Bench Handling
• Restrained Cargo Vibration
• Fragility
• Operability with Generator Power
Process Flow: DNA Purification
• All BioChipSet process steps are controlled using pneumatic pressures.
• Guanidinium/Silica filter based purification.
• Self-normalizing to delivery consistent range of DNA concentration for amplification.
Insert Swab (x5)
Cell Lysis
DNA Purification
Elute DNA
Process Flow: Rapid Multiplexed STR Amplification
• PCR uses Promega PowerPlex® 16 System.
• PCR reagents have been lyophilized for long-term stability at room temperature (>2 years).
• Rapid amplification: 31 cycles, 19.5 min.
Purified DNA Sample
Reconstitute PCR Cake
Transfer to PCR Chambers
Amplification
Process Flow: Electrophoretic Separation and Detection
Join PCR Product & Formamide
Reconstitute 5 ILS Cake/1 AL
Injection 5 Samples/1 AL
Electrophoretic Separation
Fluorescence Detection
Data Analysis and DNA Profile Generation
• Expert System software performs automated data processing & allele designation.
• Three data output files are automatically generated for database searching and
• No data review or analysis by a forensic analyst is required.
ANDE Testing
• U.S. Testing Partners: USACIL, FBI, and NIST have been performing laboratory testing for the past 12 months
• U.S. Military/Intelligence Exercise Trident Specter: deployed in maritime operations
• SOCOM: initial laboratory testing performed, currently deployed for field testing
• Australia: Successfully deployed in field exercise Q3 2013
• DHS: Immigration pilot scheduled for Q4 2013
Rapid DNA Analysis: An Enabling Technology
Rapid DNA Analysis for Human Identification is the automated generation of STR profiles (DNA fingerprints) by non-technical operators outside the laboratory. Major applications to enhance societal safety include:
• Military and Intelligence Operations– Database Enrollment– Counter Terrorism/Counter Narcotics– Maritime Operations– Enemy Combatant Identification– Casualty identification
• Homeland Security – Immigration processing– Mass Disaster Victim Identification– Borders and ports protection– Passport/document fraud
• Human trafficking prevention• Law Enforcement
– Arrestee processing – Crime laboratory support
