dr michael loughran team leader biophotonics & microfluidics research integration of silicon and...
TRANSCRIPT
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Integration of silicon and glass processing for lab on a chip development
• Dr Mike Loughran
• Tyndall National Institute
• Cork, Ireland
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Overview
• Introduction Mike Loughran, Tyndall National Institute• Lab on a chip research development• Choice of substrate and fabrication techniques• Fluidic Inter-connects• Microchip HPLC Development on silicon substrates• Capillary Gel Electrophoresis in Single Glass Microchip• Processing of Glass Microchips • Electrowetting (microfluidic transport• Encoded silicon microbead technology for lab on a chip• Integration of VSCEL (optical light source for microbead detection)• Development of customised reaction chamber for micro bead
functionalization• Chemiluminescent allergen detection• Acknowledgements
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
PhD Biotechnology Cranfield University 1995
Senior Research Fellow AIST Tsukuba Japan 2002-2004
Dr Mike Loughran Brief Introduction
JSPS Research Fellow Tokyo University of Fisheries from 1995-1997
Research Fellow University of Manchester, U.K , 1999-2001
Visiting Associate Professor Tokyo University 2001
Team Leader Biophotonics & Microfluidics Research Tyndall National Institute 2004-2006
Research Fellow Dublin City University, 1997-1999
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Lab on Chip Research Development
Micro-reactor Surface coating
Micro-sensor Optical/ Electrochemical
Micropump/valveLiquid transport
Fabrication techniques must be cost-effective Economical precise control of channel dimensions/geometry Accuratepreferably made on large scale wafers Mass production
Intrinsic Advantages of -tas lab on chip systemsHigh through put, rapid analysis, reduced reagent/sample consumption
Continuing challengesSample transport/inter-connects from bench to micro-chipReproducibility (feature size) genuine versatile platformObscure terminology “nano” or micro,
top down, bottom up
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Choice of substratePolymer micro-chips: replication technology, embossing, imprintingIn expensive, rapid proto-typing, non clean room processingSolvent in-compatibility, channels not uniform, temp defects, opaque, hydrophobic Examples Polycarbonate, PMMA, SU-8, PDMS
Silicon Micro-machiningPhoto-lithography, wet etching, dry etching, anodic bonding, dicingSystem integration: electrodes, micro-channels Not suitable for high voltage capillary electrophoresis separations silicon breaks down at voltages > 1000 V
Glass processingPlanar technology, transparent, surface properties well characterised, amenable for bio-conjugation,self assembly, facilitate high voltage separation > 50kV, clean room and no clean room processing,
Flexible processing photo-lithography, wet etching, dry etching, anodic bonding, dicing
But bonding with UV curable adhesives not always provide permanent seal
Fusion bonding (high temperature 650oC above glass transition phase) more reliable seal provided micro channel alignment can be confirmed
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Micro chip fabrication techniques
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Fabrication Feature Size
simple process for wet anisotropic channel etching with a controlled depth up to 500 nm, an accuracy of a few percent and etch roughness less than 0.5 nm
In photolithography ultraviolet light is used (typically 250 nm wavelength)
fabrication of spacings < 125 nm causes blurred features, can melt together. technical improvements enable structural resolutions ca. 70 nm in experimental setups and ca. 100 nm for mass production
Lithography technologies based on focused beams are an alternativee.g. Electron beam lithography (EBL) and focused-ion beam (FIB) lithography (feature size 10 nm) to create nanochannels for DNA separation
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Fluidic Inter-connects
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Rheodyne Valve for Fluidic Switching
Fluidic Inter-connects for microchip HPLC
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
SU-8 microfluidic Chip rapid processing
• Chip designs made from low cost acetate masks
• SU-8 lithography in the plating lab (process developed by D. Hoffman)
• Access holes drilled manually in lab 1.13
Fluidic interconnects for allergen microarray
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Preliminary packaging of allergen microarray
User-friendly chip interface
•Initial prototype chip development:
Covalent binding of PDMS on glass with oxygen plasma treatment (in plating lab)
• Plastic chip holder:
More stable solution
Holder fabricated by J. Rea
Sealing techniques (minimize dead volume)
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Integrated access holes for sample introduction
Glass microfluidic Chip processing
• Chip designs realised on chromium masks (to withstand HF etching)
•Customised Casper Process Development(
-> integration of access holes in the clean room fabrication process by deep(200 um) HF etching (better alignment, less fragile)
-> Glass channel lithography and wet etching (100 um) in a clean room environment
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Customized microchip holder
Plastic chip holder
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Packaging of HPLC Microchip
Figure 9: Problems with packaging. (a) Dead volume at the end of the optical fibre and air bubbles existing in the glue possibly cause the leakage. (b) Dead volume at the end of the fused silica capillary. (c) Blockage caused at the end of the fused silica capillary
UV glueChannel
CapillarySilicon
With UV glue
Without UV glue (Dead volume)
ChannelCapillary
Capillary end
SiliconSilicon
Capillary
Optical fibre
Dead volume Air bubble(a)
(b)
(c)
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Final Mask Design with Integrated HPLC Chips
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
HPLC Chip Fabrication A fabricated micro HPLC chip
Injection channel
Micropillar (5×5 µm)
UV detection
Injection
Separation
Figure 7
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Glass Processing:
Capillary Gel Electrophoresis
Rapid microchannel fabrication
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Separation capillary length 60 mm, diameter 500 μm, depth 300 μm.
Magnified view of self-contained stacking capillary
Length 12 mm,
diameter 1 mm,
depth of 300 μm.
Sep
aration
cap
illary
Stacking capillary
SDS/Native Capillary Gel Electrophoresis in Single Glass Microchip
Shou-Wen Tsai , Michael Loughran, Hiroaki Suzuki & Isao Karube, “Native and SDS Capillary Gel Electrophoresis of Proteins on a Single Microchip -” Electrophoresis (2004)25:494-501
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Simultaneous separation of both native and SDS marker proteins in assembled chip
Experimental conditionsconstant current of 2 mA, 10 minutes after sample pre-concentration at 50V
Simultaneous SDS Native Electrophoresis In Multiple Capillaries
Shou-Wen Tsai , Michael Loughran, Hiroaki Suzuki & Isao Karube, “Native and SDS Capillary Gel Electrophoresis of Proteins on a Single Microchip -” Electrophoresis (2004)25:494-501
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Wet etching of microfluidic structures in glass using photo-definable epoxy (SU-8) as an etching mask
• Rapid generation of microfluidic structures in glass • Using an epoxy based polymer (SU-8) as an etching mask • 49% concentrated hydrofluoric acid as the glass etchant• Generation of microfluidic structures with a maximum etch depth of 100 µm• The glass material used was Borofloat33• The wafers were 600 µm thick and non-polished (both sides)• This type of glass can also be used for anodic bonding to silicon substrates
– Fabricated microfluidic glass chips were etched for 10 minutes
– Resulting channel depth is about 70 µm
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Bonding and sealing of fabricated microfluidic glass chips
• PMMA to glass dircect bonding• PMMA/glass to glass bonding via PDMS interface layer
2 methods were applied to seal the microfluidic glass chips
Both types of bonded glass chips were tested with a fluorescent dyed liquid at different flow rates
• Maximum flow rate tested: 417 µl/s• Resutling average velocity: 15 m/s• Resulting pressure: 250kPa 36 psi• No leakage observed for both methods• No delamination observed for both
methods
Sufficient sealing for most applications
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Test of fabricated microfluidic glass chips
InletOutlet
Sequence 1 Sequence 2 Sequence 3
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Observed advantages of glass processing
• Successful implementation of a microfluidic chip manufacturing technology where microchannels are defined in glass
• Offers a very good alternative to microfluidic chips with microchannel structures defined in SU-8
• Offers alternative bonding methods avoiding the use of UV glue or SU-8 bonding techniques
• The bond is clean and of high quality in terms of uniformity and tightness
• Microfluidic glass chip is completely visualisable as both substrate and superstrate are transparent
• DNA was successfully hybridised with probe DNA prior immobilised into the channel structure
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Electrowetting
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
PDMS
W.E.
hydrophobic
hydrophobic
hydrophilic
hydrophilic
OFFOFF
Electrolyte
ONON OFFOFF ONON
hydrophilic
hydrophilic
hydrophilic
hydrophilic
ONONhydrop
hobichydrop
hobicOFFOFF
Principle of Electrowetting
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
setup
CCD camera
potentiostat
Control PC
Microfluidic chip
Electrowetting droplet transport recorded in dark room
Experimental setup
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
PDMS substrateFlow channel Reservoir
Glass substrateA.E. (Pt)R.E. (Ag/AgCl)
W.E. (Au)
WorkingElectrode
Flow channelElectrolyte
300 m
40 m
Insulating layer
PDMS electrowetting micro-fluidic chip
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
12.5
mm
23 mm
15 m
m23 mm
100 or 300 m
Wettability of glass more uniform than PDMS.Diameter of microchannel reduced from 500 to 100 mm
Electro-wetting: glass microchip fabrication
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
• Materials• Ceramic plate (Al2O3), weight, oven, pyrex glass,
• Glass cleaning
• acetone, isopropanol, deionized water, acid cleaning (H2SO4:H2O2=9:1), deionized water, drying (N2 gas)
• Ceramic plate cleaning
• Isopropanol
glass
Ceramic plate
weightSetup
Glass flow channel was bonded byFusion bonding.Alexandra helped me to completefusion bonding.
Fusion bonding: to seal glass electrowetting micro-chip
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Leakage
Sometimes glass/glass bonding is not uniform due to presence of air in capillary channel during wafer alignment.Use of a vacuum oven may minimize this problem in future
Images of glass microchip
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Encoded Silicon Microbead Technology
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Results:
Encoded Silicon Microbeads - Design and fabrication
Optically encoded
No photobleaching
Material Silicon/Silicon Oxide
High chemical stability
Design is mask programmable
Variety of shapes
Large range of sizes
High member library
Compatible with standard MEMS fabrication processes
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Multiple microbead injections: Approach I
Cartridge Development
• Loading Mechanism
• Injection Mechanism
Image sequence showing successive injection of two microbeads
System Integration - Microbead Injection
Problem: Repeatability of precise injection of individual microbeads
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Characterisation – Identification System
Implementation of optical detection system
[1]
[5]
[2]
[3]
[4]
[4] Focusing Lens - Aplanatic Doublet (f - 6 mm)
[5] Photo Detector - Silicon Phototransistor
[1] Laser Diode and Collimator - AlGaInP, 635 nm, 3 mW
[3] Beam Splitter - 50:50 Cube
[2] Aperture - 3mm
[D] Encoded microbead
1 cm
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Silicon Photo-Detector
Type B Microbead
IntegratedVCSEL
Polymer Suberstrate with Microchannel
Polymer Substrate with Cavity
Slit
Encoded Silicon Microbeads - Optical Detection
Integrated detection system: Principle of operation Detection of microbeads with through-hole barcode pattern
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
PMMA (Substrate - Cavities)
PMMA (Superstrate - Microchannels)
Casting Resin (Master - Microchannels)
Silicon (Master - Cavities)
SU-8
VCSEL Package
Process Flow - Cavities Process Flow - Microchannels
1a
2
3
1b
4
5
Encoded Silicon Microbeads - Optical Detection
Integrated detection system: Illustration of fabrication process
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Encoded Silicon Microbeads - Optical Detection
Integrated detection system: Experimental set-up and results
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
System Integration - Reaction chamber
Integrated detection system: Illustration of fabrication process
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Experimental setup:
Application – Instrumentation
Temperature Control
Electronics
Peltier Element
Temperature Feed back
loopCurrent Control
PC
Syringe Pumps
Temperature Monitoring
Syringe Control
Flow Control
Microfluidic Reaction Chamber
Temperature Sensor
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Temperature Controller – Technical Description
Temperature Controller
Electronics
Temperature Measurement
Electronics
Temperature Monitoring with
Labview
DAQ
PC
Peltier Element
Microfluidic Flowcell
Temperature Sensors
Requirements:
•Temperature range 35°C ... 45°C•Stability ± 0.5°C over a period of 1h•Control offset less than 0.2°C at 40°C
Temperature Feed back
Current Control
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
-15V
┴
+15V
DAQ
4.5V
┴
Set-pointer
Switch
PT100
PT100
Peltier
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Device simulation: Results
Application – set-up
To be finalised!
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
+15V
-15V
+5V
DAQ
Laser Diode HL4314MG
+2.7 V, 30 mA
Silicon PhototransistorSD3443
Battery
I/O Connector
DAQ Card PC
Channel 3
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Experimental Evaluation
DNA hybridisation at microbead surface in 4 x 4 array
Accepted for Lab on a chip: December 2006 in press
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Chemiluminescent Allergen Detection
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Principle of Chemiluminescent allergen detection
h h
HRP HRP
PDMS PDMS
Alergen immobilisation
Microfluidic environment
Reaction Chamber
UCBL Lyon, France
Tyndall, IrelandGlass Superstrate (SU-8 on superstrate)
PDMS Substrate
SU-8 SU-8
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Allergen deposition by piezo-electric spotter
•Matrix of alergen probes•Simultaneous detection of 48 probes•Incubation with serum of target patient
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Optimization of Fluidics by Coventor Simulation
Coventor simulations
With MEMs CFD package
Steady and dynamic flow simulations as a function of :• Microfluidic design• Chip dimension• Experimental conditions (flow rate, etc..)
Allow us to see the distribution of flow velocities, or of the filling of a liquid in the microchannel
Compared various geometries of microfluidic system and flow cell design
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Chemi-luminescent allergen detection
Interaction with UCBL (C. Marquette, K. Heyries, Lyon)
They perform: * immobilization of the proteins by spotting technique on PDMS * antigen/antibody assays with our microfluidic chip
• Assay requires uniform reagent distribution -> flow cell with optimized geometry (flow simulations using COVENTOR)
• Chip processing -> SU-8 -> Glass channels
• Need friendly chip-user interface, enabling reproducible measurements -> Chip holder
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research
Acknowledgements
• I appreciate cooperation of all members of Biophotonics and Microfluidics Research Team
• Tyndall CFF Fabrication engineers and management team for their support.
• Jenny Patterson and Intel for finance of fabrication and processing costs (EI Intel Innovation Fund)
• Wataru Satoh JSPS Research Fellowships sponsored by Japan Society for the Promotion of Science
• Dr Miloslav Pravda Dept UCC Chemistry
Dr Michael Loughran Team Leader Biophotonics & Microfluidics Research