Electrochemical Nanopatterning and Microsystems

Applied Biotechnology Innovation Centre

Electrochemical Nanopatterning and

Microsystems

Ioanis Katakis

Department of Chemical Engineering, ATIC Technology Innovation Centre, Universitat Rovira i Virgili, Tarragona, Spain

November 29, 2010

NANOJASP 2010, Barcelona


DEFINING TERMS

HOLDING EVERYTHING TOGETHER

THE TRANSDUCER

THE BIOMOLECULE(S)THE TRANSDUCTION

CHEMISTRY

MODULATING ACTIVITIES

CONTROLLING RATES OF REACTION AND MASS TRANSPORT

• The generic bioelectronic element


MODULE 1

--

MODULE 2

++ + MODULE 3

-- -

MODULE 4

Os

Os Os+ +

MODULE 5

MODULE 6...

-

-+

+

+

Os

Os

Os

+

+-

-

-

• Each module can be any of the components. You aim at flexibility and control of outcome.

MODULAR IMMOBILISATION...

• Narváez et al. Biosens Bioelectr 15:43-52 (2000)

• Narváez et al. J Electroanal Chem 430:227-33 (1997)


• Rational manipulation of bioregeneration kinetics

Au / MPS / RP / PSS / FDH

+

+Os

Os

Os

S SO3-

S SO3-

SO3-

SO3-

SO3-

ORDERED IMMOBILISATION: Multilayer self-assembled redox polyelectrolyte-FDH architecture in gold electrodes

average catalytic current = 1.04 µA RSD = 16 % n = 6

0 0.1 0.2 0.3 0.4 0.5

E / V vs Ag/AgCl

0

0.4

0.8

1.2

I / µ

AAu / MPS / RP / PSS / (B / PSS)n / FDH

S SO3-

S SO3-

+

+Os

Os

Os

SO3-

SO3-

SO3-

SO3-

SO3-

SO3-+

+

+

+

Fructose modular electrodes

…TO CONTROL PROPERTIES


SUPRAMOLECULAR ARCHITECTURES FOR COMPLEX BIOSENSING, AMPLIFICATION, AND

CATALYSIS TASKS

• Popescu et al. J Electroanal Chem 464:208-14 (1999)

Lipid

Os-Phendion-surfactant

GLDH

NAD+

Cholesterol

Substrate

Product

Dehydrogenase

NAD(P)+

NAD(P)H

Mediatorred

Mediatorox

Electrode e-

APPLY JUDICIOUSLY MOLECULAR ENGINEERING


• CONTACT AND CONTACT-LESS SPOTTING WELL DEVELOPED FOR 100+ mm RESOLUTION

• BIOMOLECULE PHOTOLITHOGRAPHY FOR HIGH DENSITY APPLICATIONS

BUT WHAT ABOUT PATTERNING?

WHAT IF WE COULD USE ELECTROCHEMICALLY-DRIVEN METHODS FOR BOTH PATTERNING AND DETECTION?


• Colloidal gold: a versatile nano/module

- - - --

---

-++

+

Electrostatic forces

Adsorption phenomena

sDative binding

“NANO” ENGINEERING SPATIAL INTELLIGENCE


• Campàs & Katakis Int J Env Anal (2004), PCT/EP2003/000262 (2002)

• Campàs & Katakis Sens. & Actuators B (2006)

gold colloidoligonucleotide

colloidal gold-oligonucleotideselective deposition

deposition

conjugation

NANOCOLLOID SYNTHESIS AND MODIFICATION FOR NANOPATTERNING AND ARTIFICIAL

INTELLIGENCE

PATTERNING BIOLOGICAL PROPERTIES


Electrochemistry on SPE

e-

0

5

10

15

20

I (nA

)

1.2V (compl)

no E (compl)

1.2V (4-mut)

no E (4-mut)

ARE THEY FUNCTIONAL?

• Concept works but high exists non-specific adsorption


GOx

GOx

GOx

GOx

GOx

• Adding properties (transduction) to nanopatterns

MOLECULAR ENGINEERING OF “NANO”...


AuAu

PEG

SH

PEG

SH

PEG

SH

PEG

SH

Thioctic acid SAM

e1 (-1.2V)

desorption of

Thioctic acid

e1 (+0.8V)

selective adsorption

of HRP-Os-Au

e2 -1.2V)

selective

desorption

The deprotection of the second electrode avoids the non-specific adsorption detected.

e2 (+1.2V)

deposition of

Gox-Os-Au

There is only a 3,4% of non-specific

response from the second electrode

H2O

2

e-

Amperometric

detection of HRP (e1) Non-specific adsorption was detected

on the second electrode

Glucos

e

e-

Amperometric detection

of GOx (e2) 5% of non-specific response was

detected from the first electrode

• Amperometric detection

of electrodeposited

biomolecules

…AND IMPROVING SELECTIVITY


SS

HO

O

OO

O

SS

HO

O

OO

O

SS

HO

O

OO

O

SS

HO

O

OO

O

SS

O

OO

O

O

O

Au

OO

O

O

O

H

SS SS

HO

O

OO

O

SS

HO

O

OO

O

SS

HO

O

OO

O

SS

HO

O

OO

O

Au

+700mV

ONE STEP FURTHER: PATTERNS AT MOLECULAR LEVEL


0.477 0.577 0.677 0.777 0.877 0.977 1.077-6-0.069x10

-6-0.019x10

-60.031x10

-60.081x10

-60.131x10

-60.181x10

-60.231x10

-60.281x10

-60.331x10

-60.381x10

-60.431x10

E / V

i / A

Peak around +0.6V

Electrochemical deprotection was nearly complete within one scan

DOES IT WORK?


0

100

200

300

400

500

600

Compoundimmobilisation

Gox(non-specif ic) Gox(AfterElectroactivation)

Fre

qu

en

cy c

han

ge(H

z)

• EQCM data shows hope (but still 30% non specificity)

IS IT SELECTIVE?


• Resist coating• First laser exposure• First biomolecule coating • Second laser exposure• Second biomolecule coating• Polyelectrolyte blocking

Laser ablation or lithography work equally well

AND YET ANOTHER METHOD OF PATTERNING


• Layer optimisation: GOX(first layer) SAOX (second layer)

Activity of glucose for every step of PE layer deposition

0

0,00000005

0,0000001

0,00000015

0,0000002

0,00000025

0,0000003

0,00000035

0,0000004

0,00000045

0 5 10 15 20 25 30

glucose

i/A

native

1st layer

2nd layer

3rd layer

GOXGOX 2h.

POS+GOX

SOX 2h.

POS+SOX

• After third layer of polyelectrolyte the response for enzyme decrease 30%.

BIOPHOTOLITHGRAPHY: CATALYSIS(1)


GOX (Max. Response 856A)

SOX (Max. Response 10 nA)

• Layer optimisation: GOX(first layer) SAOX (second layer)

0

0.0000001

0.0000002

0.0000003

0.0000004

0.0000005

0.0000006

0.0000007

0.0000008

0.0000009

0 2 4 6 8 10 12 14

substrate(mM)

i/A

Gox and SAOx response with Glucose

GOX

SAOx

GOXGOX 2h.

POS+GOX

SOX 2h.

POS+SOX



GOX (Max. Response 0A) No crosstalk.

SOX (Max. Response 101 nA)

• Layer optimisation: SAOx (first layer) GOX (second layer)

0.00E+00

1.00E-08

2.00E-08

3.00E-08

4.00E-08

5.00E-08

6.00E-08

7.00E-08

8.00E-08

9.00E-08

1.00E-07

0 5 10 15 20 25 30 35 40substrate(mM)

uA

Gox and SAOx response with Sarcosine

GOx

SAOx

SOX

GOX2h.

POS+GOX

SOX 2h.

POS+SOX



1st electrode

(Sample)

2nd

electrode

(Control)

AuAu

Deprotection of e1 with

UV

Incubation of e1 with

redox polymer and

streptavidine

Immobilisaion of

biotinylated mutated

capture probe in e2

Deprotection of e2 with UVIncubation of e2 with

redox polymer and

streptavidine

Immobilisation of

biotinylated wild capture

probe in e1

Incubation of both

electrode with the

biotinylated target

Incubation of both

electrode with

streptavidine-HRP

Amperometric detection of

both electrodes

HRP HRP

H2O2 e-

BSA blocking in

e1BSA blocking in

e2

IDE: Higher signal was obtained from the electrode with wild probe (1.7A), 0.4A was obtained from the mutated probe and 0.3 A from the control without target.

Also in CE higher signal was obtained

from the electrode with wild probe

(50nA), while 0nA was obtained from

electrode where mutated probe was

immobilised

BIOPHOTOLITHGRAPHY: HYBRIDISATION


• HCG amperometric detection through sandwich assay

1st electrode

(Sample) 2nd electrode

(Control)

AuAu

HRP

H2O2 e-

e1

deprotection

Incubation with redox

polymer and anti-HCG in

e1

BSA blocking in

e1

Incubation of HCG target

and biotinylated anti-HCG

in e1

e2 deprotectionIncubation with redox

polymer and anti-HCG in

e2

BSA blocking in

e2

Incubation of biotinylated

anti-HCG in e2 (control)

Incubation of both

electrodes with

streptavidine-HRP

Amperometric detection of HRP

IDE: lower signal was obtained comparing with DNA, however as in DNA wafers the control was lower (2nA) than the sample (40nA)

Also in CE there is a lower

response from the control,

nevertherless the signal is lower.

BIOPHOTOLITHGRAPHY: MOLECULAR RECOGNITION(1)


• T4 amperometric detection through competition assay

1st electrode

(Control)

2nd electrode

(Sample)

AuAu

H2O2 e-

e1

deprotection


polymer and BSA-T4 in e1BSA blocking in

e1

Incubation with anti-T4 in e1

(Control)e2 deprotection


polymer and BSA-T4 in e2BSA blocking in

e2

Incubation with T4 and anti-T4

in e2

Incubation of both

electrodes with anti-

Rabbit Igg-HRP

Amperometric detection of HRP

HRP

A competition assay was carried

out to detect T4. 94.2nA was

obtained from the control and

47.3nA from the sample

BIOPHOTOLITHGRAPHY: MOLECULAR RECOGNITION(2)


SHSH SH

SH

HRP

e-

H2O2

SH

SHSH

SH

A difference of 1,5µA between sampleand blank and a limit of detection of 6.31fmoles was obtained

MODULATING ACTIVITY: RECOGNITION TO SENSING


H2O2

e-

HRP

H2O2

e-

- 57% and 23% of signal displaced in colourimetric and electrochemical displacement- Fast response: 2 minutes in electrochemical displacement

AND FROM SENSING TO FACILE SENSING


Os+Os+

Os+Os+

Os+Os+

Os+Os+

Os+Os+

Os+Os+

- - --- -

+ + +++ +- - --- - }N1

+ + +++ +

+ + +++ + }N2

- - --- -

Glucose oxidase produces H2O2

Peroxidase uses H2O2 and consumes electrons at E2

Glucose oxidase produces electrons at E1

e-

e-

TOWARDS THE ULTIMATE NANOMACHINE(?): SELF POWERED, SELF PROPELLED, SELF PROPAGATING

INTEGRATING TECHNOLOGIES FOR MORE FUNCTIONS

• Pescador et al Langmuir (2008)


Using same principles for versatile microsystem operation

ELECTRODEPOSITION AS PART OF OPERATION

A

B

A

B

• Mata at al Electroch. Acta (2009)


THE TEAM

• Panagiotis Argitis

NCSR DEMOKRITOS

• Mònica Campàs

• Mònica Mir

• Srujan Dondapati

• Pablo Lozano

Universitat Rovira i Vrigili


Our work is financed by:

• MICROPROTEIN (Patterning and arraying)

• HEALTHY AIMS (Fuel cells)

• CELSITIVE (Pathogen Detection)

• CIDEM and our Clients

• URV

THE MONEY


THANK YOU FOR YOUR

ATTENTION

Documents

Electrochemical Nanopatterning and Microsystems