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.