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Transcript of Disposable molecular diagnostics: Microfluidic laboratories for the field Catherine Klapperich,...
Disposable molecular diagnostics: Microfluidic laboratories for the field
Catherine Klapperich, Ph.D.Biomedical Microdevices and Microenvironments
Laboratorywww.klapperichlab.org
Biomedical and Manufacturing Engineering Departments
Boston University3 October 2006
Microfluidics Applications• Diagnostics/Management
– Point of Care– Disease Surveillance
• Detection– Homeland Security– Fighting Force Protection
• High Throughput Screening – Drug Discovery/Development– Cell Based Assays– Research Bench Applications
• Micro-Reactions– Combinatorial Methods
• Living Tissue Arrays – Drug Development
Global Impact
• Case finding• Case management• Surveillance
24% of the current burden of disease could be averted if 80% of the population of low income countries received the following: prenatal/delivery care, family planning, treatment of TB, management of sick children and case management of STDs.
Implementation would cost $8/person per year.
Nature Reviews Microbiology 2, 231-240 (2004) DIAGNOSTICS FOR THE DEVELOPING WORLD
State of the Art• Microscopy
– Parasitic and mycobacterial infections– Requires well trained technician
• Cell Culture• EIA• Nucleic acid amplification
All require specialized equipment and/or technicians.
MicroTAS for Diagnostics
Sample Preparation
Fluidics Detection
Control and Signal Processing
Input Output
• Sample introduction• Cell sorting/separation• Mixing • Lysis• Separation/concentration• Detection• Waste stream capture
System Schematic
Detection Surfaces/Channels
AntibodiesOligosPCR
Device Design Constraints
• Inexpensive materials• Rapid prototyping• Scale up/mass production• Shelf life of 1 year or more• Ease of use• On-board reagents• Disposable• Little sample preparation off chip• Low power or no power
Materials Requirements• Optical properties
– UV transparent (for quantifying proteins, DNA and RNA)
– Transparent to excitation and detection wavelengths (488 nm, FITC)
• Thermal properties– For PCR (95 degrees Celsius)– For dimensional stability
• Surface chemistry– Hydrophilic/hydrophobic– Non-binding– Binds specific molecules– Shelf life issues
R1 R2
R1 R2
R1 R2
CH2CH
R3
H2C CHR3
n
n
m
R1 R2
n
H2
R1,R2,R3; H
TOPAS by TICONA and APEL by Mitsui
ZEONEX and ZEONOR by ZEON
AdditionPolymerization
Ring Opening Polymerization
ZEON Polymers are obtained by ring-opening metathesis polymerization (ROMP) of norbornene derivatives monomers followed by complete hydrogenation of double bonds.
Engineering Polymers for Microfluidic Diagnostic Devices
Zeonor 750R, Tg 70CZeonex 690R Tg 136C
PMMA
Polycarbonate
Polystyrene
Tg= 85-105C
Tg= 140-150C
Tg= 90-110C
Advantages of Thermoplastic Chips
• Feature size is identical to PDMS but with long term dimensional stability.
• Surface treatments are robust and do not “age” as on PDMS devices.• Permeability is low.• Thermoplastics can be purchased in grades that are certified non-
pyrogenic (do not contain DNA or RNA destructive enzymes).• The per device material cost is low.• The plastic chips can be easily manufactured in-house using rapid
prototyping techniques in production materials to test and optimize new chip layouts and chemistries quickly.
• Internal structures (filters, valves, detection patches) can be fabricated in situ by light-directed processes.
• Acrylics and cyclic polyolefins have low autofluorescence for high detection signal to noise ratios.
• Acrylics and cyclic polyolefins are transparent to UV, which enables light directed processing of internal structures and UV detection of nucleic acid concentrations and integrity through the chip.
Rapid Prototyping
Pressure and Heat Applied
DRIE
Mask
UV light
Si Wafer
Photoresist
Polymer pellets
Embossed substrate
Thermallybonded channels
• A cyclic polyolefin (Zeonex 690R) was used as chip material• Microchip fabricated by hot-embossing with a silicon master
Scale-up Fabrication
Glass or Si wafer
• Photoresist coat• Mask, expose, develop photoresist• Etch glass• Remove photoresist
Etched glass plate
Electroplate
Separate
Metal electroform
Repeat1000’s of
times
Separate
Molded plastic card
Mold or emboss
Coverandseal
Plastic cover layer
Completedfluidiccard
Light-directed Processing in Formed Channels
In Situ Filter and Column Formation
O
O
O
O
Nucleic Acid Extraction
Moving Fluid
• Pressure• Vacuum• Electroosmotic Flow
– Surface Chemistry of Channels– Simultaneous Assay and Device
Development
Immobilized Surface Chemistries for Detection
Jessica Kaufman
Arpita Bhattacharyya
Justyn Jaworski
Nathan Spencer
Dominika Kulinski
Dave Altman
Amy VanHove
Dr. Cassandra Noack
Coulter Foundation
Whitaker Foundation
CIMIT
NSF MUE
Pria Diagnostics, Inc.