microfluidics vs self assembly microfluidics vs. self ...

microfluidics vs. vs self self-assembly assembly

Andreas Manz KIST Europe, Saarbrücken, Germany KIST Seoul, Seoul South Korea Mechatronics, Saarland University, Germany

… questions like: “how is a butterfly wing manufactured?” manufactured? • microstructure, nanostructure, colour • stable t bl material t i l (chitin), ( hiti ) nott alive li • reproducibility • ease of manufacturing • low cost • … and what is the blueprint for it?

… questions like: “how is a butterfly wing manufactured?” manufactured?

•… and d what h t is i the th blueprint bl i t for f it? • … how to get from molecular biology to structure? • … how to get discrete size, structure g byy self assembly? y • … how to engineer


•… and d what h t is i the th blueprint bl i t for f it? • … how to get from molecular biology to structure? • … how to get discrete size, structure g • all 3 have identical genome


Morphidae, 170mm smallest feature 100nm


Morphidae, 170mm smallest feature 100nm


Morphidae, 170mm 100nm

… note: I am not yet speaking of engineering a microsystem like this…. this

Syrphidae 7mm Syrphidae,

what is “lab on chip” technology?

• dev device ce made de from o a substrate subs e • using clean room technology chemistry biology, biology medical use • target: chemistry, • containing channels, reactors etc • may contain t i detectors, d t t heaters, h t etc. t

why is it difficult?

• clean l room technology t h l is i expensive i • labour intensive • mistakes in layout difficult to correct y example…) p ) • ((take my

ciba-geigy, basel switzerland (now novartis, novartis solvias) 1988-1996

ciba-geigy, basel switzerland (now novartis, novartis solvias) 1988-1996

imperial college london 1996 2006 1996-2006



isas, dortmund germany 2003 2008 2003-2008

substrate materials glass‐glass glass glass

pmma‐pmma

pdms‐pdms pdms‐glass glass silicon glass‐silicon

glass‐glass silicon‐silicon

silicon‐silicon pmma‐pmma glass gold glass glass‐gold‐glass

glass‐silicon

glass‐laminate‐glass glass‐silicon‐glass ordyl multi‐layer d l l i l glass pdms‐copper

pdms‐glass

pdms‐silicon

d d pdms‐pdms

pdms‐silicon‐pdms quartz‐quartz

•

substrate materials used

integrated features porous membrane

g nothing

heaters

metal electrodes heaters porous membrane slit array outside

metal electrodes t l l t d

liquid membrane

nothing

planar waveguides x‐ray source t sensor phase quides

•

integrated featurs, like metal electrodes, heaters, membranes etc

topology of channels non‐binary branching

well

binary branching structure tree, spider single channel 1 loop central ch array, tree central bed, ch around

central bed, ch around tree, spider

central ch array, tree

binary branching structure non‐binary branching well central chamber, frit, tree

1 loop

single channel

network central chamber, single ch

•

topology

•

p tree, loop, p network, etc spider,

interfacing type

large holes, thick pdms cover flat plastic plates

large holes, thick glass cover

eppendorf pipets open eppendorf pipets, open fused silica tubing eppendorf pipets, open

flat metal plates

don't know, not used plastic tubing ‐ glue flat metal plates large holes, thick glass cover

plastic tubing ‐ glue

flat plastic plates fused silica tubing

don't know, not used

•

interfacing type

•

p to world interface”) (“chip

large holes thick pdms cover large holes, thick pdms cover

application area

sample p p g prep pumping application

separation

basics

d t ti detection biology

separation

reaction biology basics application

reaction

pumping sample prep

detection

•

what is the chip used for?

commercializations

commercial attempt no attempt

•

commercial attempt no attempt

How many chips were in direct line to commercialization?

take the best example

• capillary ill electrophoresis l t h i • scaling: 100x smaller (length) • time to result: < 10,000x faster g RNA or DNA analysis y • targets

•

capillary electrophoresis, flow injection, electrochemical detection

•

glass – g g glass chip, p design g 1989, fab 1989 mettler imt switzerland

•

manz, fettinger, lüdi, widmer, svs bulletin 5, 4-10, 1990

•

capillary electrophoresis, injection, electrochemical detection

•


•

manz, harrison, fettinger, verpoorte, lüdi, widmer, proc. transducers 1991 san francisco, 939-941, 1991

•

capillary electrophoresis, injection

•


•

effenhauser, manz, widmer, anal.chem. 65, 2637-2642, 1993

•

synchronised cyclic capillary electrophoresis, injection

•


•

burggraf, manz, effenhauser, verpoorte, de rooij, widmer, j.high resolut.chromatogr. 16, 594-596, 1993

•

2d capillary electrophoresis, injection

•

quartz – q q quartz chip, p design g 1996, fab 1997 imm mainz ggermany y

•

becker, lowack, manz, j.micromech.microeng. 8, 24-28, 1998

•

capillary electrophoresis, parallel processing, injection

•

glass – g g glass chip, p design g 1996, fab 1996 caliper p ltd. california usa

•

manz, becker, proc. transducers 1997 chicago, 915-918, 1997

•


•


•

manz, becker, proc. transducers 1997 chicago, 915-918, 1997

•


•


•

manz, bousse, unpublished

(patent filing 2002)

… and some results

•p proof oo oof principle p cpe • high speed separation • commercial product • market needs just 2x faster electrophoresis • (…. ( hhow di disappointing!) i ti !)

capillary electrophoresis results flow injection results electrochemical results missing

•

capillary electrophoresis, flow injection, electrochemical detection

•

g lüdi, widmer, svs bulletin 5, 4-10, 1990 manz, fettinger,

•


•

g verpoorte, p lüdi, widmer, p proc. transducers manz, harrison, fettinger, 1991 san francisco, 939-941, 1991

•

capillary electrophoresis, small molecules, fluorescence

•

p fettinger, g p paulus, lüdi, widmer, manz, harrison, verpoorte, j.chromatogr. 593, 253-258, 1992

•

harrison, manz, fan, lüdi, widmer, anal.chem. 64, 1926-1932, 1992

•

capillary electrophoresis, amino acids

•


•

capillary electrophoresis, phosphorothioate oligomers

•

paulus, manz, widmer, anal.chem. 66, 2949-2953, 1994 effenhauser, p

•

capillary electrophoresis, fractionation, phosphorothioate oligomers

•


•

synchronised cyclic capillary electrophoresis, injection

•

burggraf, manz, effenhauser, verpoorte, de rooij, widmer, j.high resolut.chromatogr. 16, 594-596, 1993

•

burggraf, manz, verpoorte, effenhauser , widmer, de rooij, sens actuators b20 103-110 1994

•

synchronised cyclic MEKC or capillary electrophoresis, injection

•

von heeren, verpoorte, manz, thormann, anal.chem. 68, 2044-2053, 1996

•

synchronised cyclic MEKC or CE, theophylline immunoassay

•


•

synchronised cyclic MEKC, human urine, derivatized with FITC

•


•

synchronised cyclic gel electrophoresis, amino acids

•

von heeren, verpoorte, manz, thormann, j.microcolumn separations 8, 373-381, 1996

•

synchronised cyclic gel electrophoresis, phosphorothioate oligonucleotides T2-T10

•

von heeren, verpoorte, manz, thormann, j.microcolumn separations 8, 373-381, 1996

•

label-free carbohydrate detection, holographic optical element

•

burggraf, krattiger, de mello, de rooij, manz, the analyst 123, 14431447, 1998

capillary electrophoresis results missing

•


•

g 8, 24-28, 1998 becker, lowack, manz, jj.micromech.microeng.

•


•

proc. transducers 1997 chicago, g 915-918, 1997 manz, becker, p

•


•


•


•


•


•


•


•

p manz, bousse, unpublished

•

presented first at ISPPA, tomakomai, japan 1998

•


•


•


•


•


•


•


•


•


… everything quite an effort …

• see seekingg alternatives e ves • particulary for manufacturing • looking at examples in nature • structured approach • self lf assembly bl

starting very simple

• 3 phase system assembly”, energy driven • “self self assembly • a droplet (just that)

Virtual Reaction Chamber Key properties – Water-based sample encapsulated by oil – (RT) PCR conducted on a glass cover slip – Micromachined heater/sensor are separated from the sample

PCR Sample

Oi l B

– Cover slip is disposable – Small sample volume makes system very fast

Mirror reflection

VRC details LENGTH

Key properties – VRC with glass placed on a micromachined silicon – Heater integrated with temperature sensor – Heating rate: thermal mass, available power with PID control – Cooling rate:  (thermal time constant)

H   ; P  GT G

LINK

SENSOR

HEATER

Ultrafast VRC, 650 K/s!!! From room temperature to 150 oC in 0.2 s!! 180

700

o

Tempera ature ( C)

140

600

120 100 500 80 60 400

40 20

0

5

10

Time (s)

15

20

Fluoresc cence (mV V)

160

Avian Influenza Virus Detection by RT-PCR Key properties Tempera ature (V)

0.6

• SYBR-Green Real-Time RTPCR

2 1

• Melting Curve Analysis

0

• 8 minutes utes for o RNA N detection detect o

-2

Fluoresc cence (mV)

10

100

Critical Threshold 22.3

-3

10

50

0 -4

0

10

20

Cycle Number

30

40

10

Flu uorescence e (V)

150

Differential Flu uorescence (V/cycle)

PCR

-1

0.3

Hot Start

-2

Virus irusD Detected

-3

RT

-4 0.0

0

2

4

6

Time(min)

8

10

-5 12

Palm-sized PCR

sample preparation 1) disruption of spores by superheating for fast DNA extraction 2) protein and peptide decomposition by superheating

63

superheating solvent is at a temperature higher than boiling point PCR without boiling! Oi

Sample

l B

mirror reflection

experiment no boiling of aqueous solutions at 240 °C for more than 30 min!!! limited by y thermal decomposition p of surroundingg oil

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 - aging - heat: dry: 150 °C ca. 1 h boiling: ca ca. 5 h electron microscope cross-section 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)

Z i Axiotron Zeiss A i 2, 2 1500 magnification ifi i 66

B. subtilis purified spores after SUPERHEATING microscope image of Bacillus subtilis spores after contrast staining (spores: blue)

Z i Axiotron Zeiss A i 2, 2 1500 magnification ifi i 67

spore disruption

Fluorescence inttensity

destruction of spores by superheating 10

1

10

0

10

-1

10

-2

p o s itiv e c o n tr o l n e g a tiv e c o n tr o l s p o r e s o lu tio n s p o r e s a fte r p r e tr e a tm e n t s p o r e s a fte r s u p e r h e a tin g

0

5

10

15

20

25

30

35

40

C y c le N u m b e r 68

protein- and peptide protein peptidep by y superheating p g decomposition for peptide mass fingerprinting

69

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

% Intensity

100 90 80 70 60 50 40 30 20 10

100 90 80 70 60 50 40 30 20 10

640.8

1232.6

1824.4

Mass (m/z)

2416.2

3008.0

640 8 640.8

1824 4 1824.4

2932.6687 2 2.0E+4

2835.6299 2884.6804 2915.7068

2932.6814 2

3008 0 3008.0

2682.5625 2724.5742

1691.1270

1635.0637

1467.3354 1498.2889 1539.3729

2416 2 2416.2

Superheating p g to 130 °C for 20 s

Mass (m/z)

2885.6414

2724.5750

1824 4 1824.4

Mass (m/z)

1363.2769 1232 6 1232.6

Superheating to 130 °C for 10 s 1635.0529

1467.3292 1475.3208 1232 6 1232.6

978.8941

640 8 640.8

379.0871

71.3642

49 0 49.0

978.5501

360.343

213.122

% Intensity

5354.2

49 0 49.0

% Inte ensity

2628.2

1467.313

No heatingg

49.0

100 90 80 70 60 50 40 30 20 10

2932.645

peptide decomposition by superheating

2416 2 2416.2

3008 0 3008.0

71

how about a challenge? • manufacture an object which hi h you can hhold ld by b hand h d • from smaller parts which you cannot hold by hand • byy self assemblyy • by structured approach

Atoms

Organelle

Smooth muscle cell

Molecule 2 Cellular level Cells are made up of molecules.

1 Chemical level At Atoms combine bi tto fform molecules. l l

Smooth muscle tissue Cardiovascular system Heart Blood vessels

3 Tissue level Tissues consist of similar types of cells. Blood vessel (organ) Smooth muscle tissue Connective tissue

Epithelial tissue 4 Organ level Organs are made up of different types of tissues. 6 Organism level The human organism is made up of many organ systems.

system level g y 5 Organ Organ systems consist of different organs that work together closely.

hierarchical assembly y

self assembly

Y.-H. Y H Jhang et al., al Organic Electronics, Electronics 13(10), pp. 1865-1872, 2012

K. Hosokawa, I. Shimoyama, and H. Miura, S & Actuators A t t A 57, 57 pp. 117 125 1996 Sensors A, 117-125,

S. A. Stauth, C. J. Morris, and B. A. Parviz,in Evolvable Hardware 2004, Seattle, WA, 2004

T. L. Breen et al., Science, 284, pp. 948-951, 1999

self assembly y

S. E. Chung et al., Nature Materials, 5, pp. 1147, 2008

S. A. S A Stauth St th and dB B. A A. Parviz, P i PNAS, PNAS 103(38), 103(38) pp. 13922-13927, 2006

C. Lin, Y. Liu, and H. Yan, biochemistry, 48(8), pp. 1663-1674, 2009

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 • assembly at fluidic interface

lateral capillary force

capillary force Su ace Surface tension (γ) Meniscus Contact angle Tripod (θc)

Air

Water

Su ace Surface tension Meniscus

capillary attraction

As approaching each other, other the contact angle is decreased and laterally attractive capillary force is increased P. Singh et al., Soft Matter, 2010, 6, 4310-4325

capillary attraction Capillary attraction between hydrophobic & floating material

http://www.youtube.com/watch?v=TAY6RcJPEHY

size effect

In order to increase Bond number, higher density, larger size, and weaker surface tension of floating material and medium are necessary necessary.

examples of LOGIC for self-assembly

B’

B can bind to

examples of LOGIC for self-assembly

symmetric type • ANTHRACENE • Six different tripods are needed for this assembly.

B

B A’

A B

C A B

B

C

A’ B

C

B

C

2x B

B

(upset B patterns are C same!) !)

B

B

B’

B’

C X and X’ are pairs p

C

C

C

2x

2x C

C’

C’

less symmetric type • PHENANTHRENE • 13 different tripods are needed for this assembly.

B

B A’

A B

C A B

B

B

D

A’ B

D

B

C

2x B

B

B C

B

D

B’ B B B’

C

D

D’ D

D’

E

E

A’ A B

B

B B

C A’ A

D ’

E

B

B B

B

D F E’ B’ B’ B’

B

F D’

D’

C

F C F’

D

C D D’

C

C’

C’

design and fabrication of assembling elements

our choice • Tripod: Plastic (SU-8) • Interface: water/air 3) Densitof(g/cm Density Yo 500 ng’s mod modulus(GPa) • Material The dimension tripods: L~ LYoung’s μml s(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 SU 8 Tripods

Coating omnicoat and SU-8 2050 on the wafer

Curing at RT and patterning

Releasing the tripods by dipping in the Remover PG

1 Omnicoat is used as releasing 1. agent of SU-8 microstructure. 2. The stress of the structure should be minimized (RT curing, no sudden thermal-process) 3 The 3. Th way to t gather th the th tripods ti d without stacking each other is necessary

Filtration for obtaining SU-8 tripods

first design for tripods

first design for tripods

process flow

1. Fabricated SU-8 pattern

2. Diced sample

3. Release of tripods from the wafer

6. Filtration

7. Washing with D.I. water thoroughly

8. Vacuumdrying of the filter paper

4. Placement of the filter paper on the filter

5. Configuration of the filtration system with vacuum pump

9. Observation with microscope

10. The petridish with floating tripod elements

fabricated tripods

Fabricated SU-8 tripod on the wafer

Released SU-8 tripod

assembly results

ideal dimer formation

observed dimer formation

assembled tripods

rectangular t l tip ti case

elimination of local minimum - Elimination of local minimum

A

B

- Elimination of local minimum - Round tip p for minimizing the interacting area - Sliding i i gradient i

results (A type)

results (B type)

tripods

tripods

tripods

tripods

snapshots

t=0 s

t=0.25 s

t=0.5 s

t=0.75 s

t=0.8 s

t=1 s

smaller tripods

Th attractive tt ti force f i nott The is strong enough to make them assembled because the smaller size leads to smaller bond number and interfacial deformation

… lipid extrusions …

• lipid li id tubules t b l • reproducible • um size • ((lifetime limited))

vesicle p production PDMS

Si

PDMS

vesicle production p

PDMS

Si 2 µm

PDMS

100 µm

vesicle production p flow direction

100 µm

side view

100 µm

side view fluorescence image

P.S.Dittrich, M.Heule, P.Renaud, A.Manz Lab Chip 6, 6 488 488-493 493 (2006)

formation of vesicle tubes

Formation of helices

50 µm

P.S.Dittrich, M.Heule, P.Renaud, A.Manz Lab Chip 6, 488-493 (2006)

… ongoing work …

• spontaneous t extrusions t i • parallel extrusions • tubing, cilia, large surface materials • soft materials

CONCLUSION

CONCLUSION

• biomimetic microfabrication may be very interesting for manufacturing ill curiosity i i driven, di l stage • still very early • concepts for selective hierarchical assembly y needed

acknowledgement k l d t Leon Abelmann, PhD, Professor Pavel Neuzil, PhD Matthias Altmeyer, Altmeyer PhD Eric Castro, PhD Adam Pribylka V Vanessa Al Almeida id Per Arvid Loethman Seung Jae Lee Mi Jang Himani Sharma Jukyung Park Christian Ahrberg Tim Mehlhorn Camilaa Madeira Ca ade a Campos Ca pos Marc Pichel

In Korea: Tae Song Kim, KIST Seoul, Korea Seoul Korea Seungwon Jung , KIST Seoul, Min Cheol Park , KIST Seoul, Korea Pavithra Sukumar , KIST Seoul, Korea