microfluidics vs self assembly microfluidics vs. self-assembly Andreas Manz KIST Europe, Saarbrücken, Germany KIST Seoul South Korea KIST Seoul, South Korea Mechatronics, Saarland University, Germany
… questions like: “how is a butterfly wing manufactured?” wing manufactured? • microstructure, nanostructure, colour t bl t i l ( hiti ) t li • stable material (chitin), not alive • reproducibility • ease of manufacturing • low cost • … and what is the blueprint for it?
… questions like: “how is a butterfly wing manufactured?” wing manufactured? d h t i th bl i t f it? •… and what is the blueprint for it? • … how to get from molecular biology to structure? • … how to get discrete size, structure • … how to engineer by self assembly? g y y
… questions like: “how is a butterfly wing manufactured?” wing manufactured? d h t i th bl i t f it? •… and what is the blueprint for it? • … how to get from molecular biology to structure? • … how to get discrete size, structure • all 3 have identical genome g
what is “lab on chip” technology? • device made from a substrate dev ce de o subs e • using clean room technology • target: chemistry biology medical use • target: chemistry, biology, medical use • containing channels, reactors etc t i d t t h t t • may contain detectors, heaters, etc.
why is it difficult? l t h l i i • clean room technology is expensive • labour intensive • mistakes in layout difficult to correct • (take my example…) ( y p )
integrated features heaters porous membrane nothing t l l t d heaters g metal electrodes heaters porous membrane slit array outside metal electrodes slit array outside liquid membrane planar waveguides x‐ray source nothing t sensor phase quides • integrated featurs, like metal electrodes, heaters, membranes etc
topology of channels binary branching structure non‐binary branching well tree, spider single channel central bed, ch around single channel 1 loop central ch array, tree central bed, ch around tree, spider central ch array, tree binary branching structure non‐binary branching well central chamber, frit, tree single channel 1 loop network central chamber, single ch • topology • spider, tree, loop, network, etc p p
interfacing type flat plastic plates large holes, thick pdms cover large holes, thick glass cover eppendorf pipets open eppendorf pipets, open flat metal plates eppendorf pipets, open fused silica tubing don't know, not used plastic tubing ‐ glue fused silica tubing plastic tubing ‐ glue flat metal plates large holes, thick glass cover flat plastic plates large holes thick pdms cover don't know, not used large holes, thick pdms cover • interfacing type • (“chip to world interface”) p
application area pumping sample prep basics application p p g separation d t ti separation biology detection reaction biology reaction basics application pumping pumping sample prep detection • what is the chip used for?
take the best example ill l t h i • capillary electrophoresis • scaling: 100x smaller (length) • time to result: < 10,000x faster • targets RNA or DNA analysis g y
• capillary electrophoresis, parallel processing, injection • glass – glass chip, design 1996, fab 1996 caliper ltd. california usa g g p g p • manz, becker, proc. transducers 1997 chicago, 915-918, 1997
• capillary electrophoresis, parallel processing, injection • glass – glass chip, design 1996, fab 1996 caliper ltd. california usa g g p g p • manz, becker, proc. transducers 1997 chicago, 915-918, 1997
• 2d capillary electrophoresis, injection • glass – glass chip, design 1996, fab 1996 caliper ltd. california usa g g p g p • manz, bousse, unpublished (patent filing 2002)
… and some results • proof of principle p oo o p c p e • high speed separation • commercial product • commercial product • market needs just 2x faster electrophoresis ( h di i ti !) • (…. how disappointing!)
… everything quite an effort … • seeking alternatives see g e ves • particulary for manufacturing • looking at examples in nature • looking at examples in nature • structured approach lf bl • self assembly
Virtual Reaction Chamber Key properties – Water-based sample encapsulated by oil – (RT) PCR conducted on a PCR Oi glass cover slip – Micromachined heater/sensor Sample Oi l B are separated from the sample – Cover slip is disposable – Small sample volume makes system very fast Mirror reflection
VRC details LENGTH HEATER SENSOR Key properties – VRC with glass placed on a LENGTH LINK SENSOR micromachined silicon – Heater integrated with LINK temperature sensor – Heating rate: thermal mass, available power with PID control – Cooling rate: (thermal time constant) H T G P G H ;
Avian Influenza Virus Detection by RT-PCR Key properties • SYBR-Green Real-Time RT- PCR 0 .6 2 ature (V) • Melting Curve Analysis • 8 minutes for RNA detection 0 1 Tempera 8 utes o N detect o 0 .3 -2 -1 e (V) V irusD etected H ot S tart P C R 100 150 10-2 cence (mV) uorescence (V/cycle) -3 uorescence V irus D etected R T 0 50 10-3 Fluoresc Differential Flu Critical Threshold 22.3 0 2 4 6 8 1 0 12 0 .0 -5 -4 Flu 0 10 20 30 40 10-4 Cycle Number T im e (m in)
sample preparation 1) disruption of spores by superheating for fast DNA extraction fast DNA extraction 2) protein and peptide decomposition by 2) protein and peptide decomposition by superheating 63
superheating solvent is at a temperature higher than boiling point without boiling! PCR Oi without boiling! Sample Oi l B experiment mirror reflection no boiling of aqueous solutions at 240 °C for more than 30 min!!! limited by thermal decomposition of surrounding oil y p g temperature x exposure time = applied energy 64
Bacillus spore disruption by superheating spores of bacteria are highly resistance against: - dryness y - toxic substances - other aggressive substances substances - aging - heat: dry: 150 °C ca. 1 h boiling: ca 5 h boiling: ca. 5 h electron microscope cross section of a spore of Bacillus electron microscope cross-section of a spore of Bacillus subtilis, showing the cortex and coat layers surrounding the core (dark central area). spore is 1.2 µm across. (Picture: S. Pankratz, Berkeley University of California) 65
B. subtilis purified spores microscope image of Bacillus subtilis spores after contrast staining (spores: blue) contrast staining (spores: blue) Z i A i 2 1500 ifi i 66 Zeiss Axiotron 2, 1500 magnification
B. subtilis purified spores after SUPERHEATING microscope image of Bacillus subtilis spores after contrast staining (spores: blue) contrast staining (spores: blue) Z i A i 2 1500 ifi i 67 Zeiss Axiotron 2, 1500 magnification
spore disruption destruction of spores by superheating 1 0 1 p o s itiv e c o n tro l n e g a tiv e c o n tro l 1 0 0 1 0 tensity s p o re s o lu tio n s p o re s a fte r p re tre a tm e n t s p o re s a fte r s u p e rh e a tin g 1 0 -1 1 0 scence int 1 0 -2 1 0 Fluore 0 5 1 0 1 5 2 0 2 5 3 0 3 5 4 0 1 0 C yc le N u m b e r 68
start with “easy” samples: ACTH • adrenocorticotropic hormone (fragment 1-24) • molecular weight 2933.44 Da • ACTH is a biomarker for cellular stress, infections, cancer (metastases!), activates G proteins… 70
how about a challenge? • manufacture an object hi h h ld b h d which you can hold by hand • from smaller parts which you cannot hold by hand • by self assembly y y • by structured approach
Organelle Atoms Smooth muscle cell Molecule Atoms Chemical level At bi t f l l Cellular level Cells are made up of molecules. 1 2 Cardiovascular system Atoms combine to form molecules. Tissue level Smooth muscle tissue 3 system Tissues consist of similar types of cells. Blood vessel (organ) Heart Blood vessels Connective tissue Smooth muscle tissue Organ level Epithelial tissue 4 Organ level Organs are made up of different types of tissues. Organ system level Organism level 4 5 6 g y Organ systems consist of different organs that work together closely. Organism level The human organism is made up of many organ systems. 6
self assembly self assembly S. A. Stauth, C. J. Morris, and B. A. Parviz,in Evolvable Hardware 2004, Seattle, WA, 2004 Y H Jhang et al Organic Electronics Y.-H. Jhang et al., Organic Electronics, 13(10), pp. 1865-1872, 2012 K. Hosokawa, I. Shimoyama, and H. Miura, S & A t t A 57 117 125 1996 T. L. Breen et al., Science, 284, pp. 948-951, Sensors & Actuators A, 57, pp. 117-125, 1996 1999
self assembly y S. E. Chung et al., Nature Materials, 5, pp. 1147, 2008 S A St th d B A P i PNAS 103(38) C. Lin, Y. Liu, and H. Yan, biochemistry, 48(8), pp. 1663-1674, 2009 S. A. Stauth and B. A. Parviz, PNAS, 103(38), pp. 13922-13927, 2006
concept • the use of hard material • achievement of asymmetric pattern by logical sequence • morphology-based assembly (non chemical functionalization) ) • capillary force as driving force • tripods as building blocks • tripods as building blocks • assembly at fluidic interface
capillary attraction As approaching each other the contact angle is decreased P. Singh et al., Soft Matter, 2010, 6, 4310-4325 As approaching each other, the contact angle is decreased and laterally attractive capillary force is increased
size effect In order to increase Bond number, higher density, larger size, and weaker surface tension of floating material and medium are necessary necessary.
our choice • Tripod: Plastic (SU-8) • Interface: water/air • The dimension of tripods: L 500 μm Material Densit (g/cm3) Yo ng’s mod l s(GPa) • The dimension of tripods: L~ 500 μm Material Density (g/cm3) Young’s modulus(GPa) Silicon 2.33 130-188 SU-8 1.19 4.02 PDMS 0.965 0.0018 Polyimide 1.43 3.2
Fabrication Procedure of SU 8 Tripods 1 Omnicoat is used as releasing Fabrication Procedure of SU-8 Tripods 1. Omnicoat is used as releasing agent of SU-8 microstructure. 2. The stress of the structure should Coating omnicoat and SU-8 2050 on the wafer be minimized (RT curing, no sudden thermal-process) 3 Th t th th t i d 3. The way to gather the tripods without stacking each other is necessary Curing at RT and patterning necessary patterning Filtration for obtaining SU-8 tripods Releasing the tripods by dipping in the Remover PG
process flow process flow 1. Fabricated SU-8 pattern 2. Diced sample 3. Release of tripods from the wafer 4. Placement of the filter paper on the filter 5. Configuration of the filtration system with vacuum pump 6. Filtration 7. Washing with D.I. water 8. Vacuum- drying of the 9. Observation with microscope 10. The petridish with floating tripod thoroughly filter paper elements
elimination of local minimum elimination of local minimum - - Elimination of local Elimination of local Elimination of local Elimination of local minimum minimum A Elimination of local Elimination of local - - Elimination of local Elimination of local minimum minimum - - Round tip for Round tip for p p minimizing the minimizing the interacting area interacting area i i i i i i - - Sliding gradient Sliding gradient B
smaller tripods smaller tripods Th tt ti f i t The attractive force is not strong enough to make them assembled because them assembled because the smaller size leads to smaller bond number and interfacial deformation
CONCLUSION CONCLUSION • biomimetic microfabrication may be very interesting for manufacturing ill i i d i l • still curiosity driven, very early stage • concepts for selective hierarchical • concepts for selective hierarchical assembly needed y
k l d t acknowledgement Leon Abelmann, PhD, Professor Pavel Neuzil, PhD Matthias Altmeyer PhD Matthias Altmeyer, PhD Eric Castro, PhD Adam Pribylka V Al id In Korea: Vanessa Almeida Per Arvid Loethman Seung Jae Lee Tae Song Kim, KIST Seoul, Korea Seungwon Jung KIST Seoul Korea Mi Jang Himani Sharma Jukyung Park Seungwon Jung , KIST Seoul, Korea Min Cheol Park , KIST Seoul, Korea Pavithra Sukumar , KIST Seoul, Korea Christian Ahrberg Tim Mehlhorn Camila Madeira Campos Ca a ade a Ca pos Marc Pichel
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