Lab-on-a-chip...Funded by the European Union - GA 737089 Kick-Off Meeting, Barcelona, Jan 17-18, 2017 1 ChipScope project kick-off meeting Lab-on-a-chip Rudolf Heer, [email protected],

Funded by the European Union - GA 737089 1Kick-Off Meeting, Barcelona, Jan 17-18, 2017

ChipScope project kick-off meeting

Lab-on-a-chip

Rudolf Heer, [email protected], AIT


Austrias biggest non-university research institute in numbers

Owners: 50.46% Republic of Austria

BMVIT Federal Ministry for Transport, Innovation and Technology

49.54% Federation of Austrian Industries

Employees: 1100 plus >250 on contract basis, thereof >100 PhD students

Financial Goal: 30% Cooperative Research

30% Contract Research

40% Basic Funding

Total Operating Income: >120 Mio. Euro



Diagnostic

Biosensors

Diagnostic

Biosensors

Business Unit Molecular Diagnostics

Saliva Diagnostics

Biomarker

Development

Assay

Development

Systems Integration

Point-of-Care

Bioinformatics


Diagnostic biosensor development


Biosensing- targets & physical transduction principles

Peptides(e.g. hormones)

Proteins(e.g. antibodies,

enzymes, hormones)

Nucleic acids(DNA, RNA,

mRNA, miRNA)

Viruses, cells

Biological targets

Waveguides

Surface plasmonresonance (SPR)

Fluorescence

Chemo-luminescence

White light interferometry

Magneticnanoparticles

Magneto-resistance(GMR, TMR)

Micro-nuclear magneticresonance

Colorimetric(lateral flow devices)

Quartz crystal microbalance

Surfaceacoustic waves

Amperometry

Voltammetry

Impedancespectroscopy Cantilevers

Optical MagneticElectro-

chemicalMassbased

Sample liquid

Signal readout& processing

Transducer

Receptor surface

Biosensor

Display of results

Waveguides Magneticnanoparticles


αφ

Diagnostic biosensor development @ AIT


Photonic simulation cluster: 256 cores with 488GB RAM

1TB high performance SSD array and

5TB raid-5 HDD storage, automated backup

• 2D/3D finite difference time domain simulation packages

• variational mode matching tools for eigenmode calculation

• Floquet-Bloch solver for simulating waveguide grating couplers

Temperature controlled sample holder

and programmable fluid control system

with five fluid reservoirs

Fully equipped set-up for waveguide device characterization and biosensing experiments

3D FDTD simulation of photonic wire Bragg reflector

Photonic design & characterization


Mach Zehnder interferometer waveguide biosensor

binding events change phase

velocity of light in sensing arm

Concentration

Intensity

Biosensitive layer on top of waveguide provides selective binding and enrichment of biomolecules on sensor surface

substrate ns

core layer nwg

waveguide side view

sensing layer nslLight

intensity

H

wwaveguide front view

analyte


Human IgG detection in serum on optical waveguides

analyte & 2nd Ab

serum

analyte only

serum

Human IgG (hIgG) detection in serumLimit of detection with secondary antibodies:0.1nM (≙15ng/ml)


Miniaturization: optoelectronic integration

optoelectronic layerSi-photodiodes

silicon dioxid 1.5-2 µm

Si3N4 waveguidesfabricated with CMOS compatibleback-end process

input grating output grating

optical sensor element

silicon substrateTSV

contactpad

=850nm


Magnetic biosensors

Based on magnetoresistance (GMR, TMR)

Electronic readout: Stray field of magnetized markers alters resistance

magnetizing field

Si/SiO2

SiO2

APTES

sensor

Hex




0 10 20 30 40 50 600,00

0,05

0,10

0,15

0,20

0,25

0,30

GMR data

exp. fit function

GM

R [

%]

sensor surface coverage [%]

blank level

Magnetic biosensors



-300 -200 -100 0 100 200 300

11,20

11,25

11,30

11,35

11,40

11,45

11,50

11,55

11,60

11,65 parallel

perpendicular

to annealing direction

res

ista

nc

e (

Oh

m)

magnetic field (Oe)-300 -200 -100 0 100 200 300

11,20

11,25

11,30

11,35

11,40

11,45

11,50

11,55

11,60

11,65 parallel

perpendicular


res

ista

nc

e (

Oh

m)

magnetic field (Oe)

-300 -200 -100 0 100 200 300

11,20

11,25

11,30

11,35

11,40

11,45

11,50

11,55

11,60

11,65 parallel

perpendicular


resis

tan

ce (

Oh

m)

magnetic field (Oe)

2.5 nm MgO

Magnetic biosensors




Easily integrated, no background signal

Flexible: average signal across certain area ↔ single marker resolution

Example: CMOS-integrated TMR biochip (mnt-era.net project MinoLab)

Magnetic biosensors




Easily integrated, no background signal

Flexible: average signal across certain area ↔ single marker resolution

Example: CMOS-integrated TMR biochip (mnt-era.net project MinoLab)

Magnetic biosensors


Tunneling magnetoresistance (TMR) biosensor

Detection of individual magnetic beads by CMOS-integrated TMR sensing cells

10 µm

Wheatstone bridge sensor with magnetic beads at surface (red bold curve) shows clearly distinguishable characteristics to out-of-plane magnetizing fields

Results of mnt-era.net project MinoLab & FP7 project e-brains

-200 -150 -100 -50 0 50 100 150 200 250

-50

-40

-30

-20

-10

0

10

20

30

40

dig

ita

l o

utp

ut sig

na

l (o

ffse

t b

y a

ve

rag

e)

out-of-plane magnetizing field (Oe)

sensor with magneticbeads on top

München


Magnetic nanoparticle-based molecular biosensing

Mix & measure detection principle – homogeneous assay

+sample

cuvette


Analyte molecule

specifically binds to

functionalized nanoparticle



On analyte

binding, a

measureable

physical property

of the nanoparticle

changes







Nanoprobes: noble-metal shell coated Co-nanorods

Co-nanorods synthesized by

organometallic approach

• Particles obtained from LPCNO-

INSA, Toulouse, France

• Single crystalline hcp Co

• Usually dissolved in organic

solvents with HDA as ligand

magnetic core length 40..200 nm

magnetic core Ø 6 nm

Nobel metal coating procedure

• Interlayer on Co-nanorods reduces

difference in surface energies

between Co and Pt/Au

• Overgrowth of Au on Pt-coated Co-

nanorods



Diagnostic prototype instrument

-200 -100 0 100 200

2.18

2.22

2.26

2.30

2.34

time (ms)

-2

-1

0

1

2

φ

small coil driving voltage (V)

photodetectorsignal (V)

power supply unitMeasurement unit & notebook

laser diode

photodetector

coil pairs

PC audio amplifier

Lock-in amplifier

sample


0 1 2 3 4 5 6 7 8

0

2

4

6

8

10

12

14

ph

ase

lag

diffe

ren

ce

(°)

sHER2 concentration (nM)

HER2 assay results

• Buffer: 10 mM HEPES, 150 mM NaCl, 0.05 v% Tween 20, pH 7.4• Nanoparticle concentration: 10 pM• Rotating magnetic field: 1 kHz frequency, 10 mT amplitude

analyte & 2nd Ab

10% serum

analyte only

buffer

analyte & 2nd Ab

10% saliva

analyte & 2nd Ab

buffer

Extrapolated limit of detection:2 nM in undiluted saliva


Electrochemical detection of proteins

Detection of the hepatitis antibody

Au surface

immobilized antigen

blocking

surface functionalizationNyquist plots for different hepatitis B antibody concentrations


Fully printed electrochemical sensors


Impedance Sensing on Adherent Cell Cultures

• Long-term monitoring for toxicological investigations• (General) Monitoring technique for cell research?

• Impedance measurement is a possible way• Wireless approach to give operators in a lab a maximum freedom of

movement and avoids error-prone cable connections



3,5 h


8 h



24 h



48 h




Adherent cells attach on bottom surface

Cell membrane with cytoplasm has higher impedance than surrounding (ionic conductive) culture media

Electrodes integrated in bottom surface

Cell adhesion

Settling Adhesion/Spreading



Sensor Device Components



Assembled sensor device



RF-based system – Base station

Antenna coil

to PC

Wireless transferof Energy, Data,and Commands

Base station

µC

MUX

RF frequencygenerator

RFID receiver

Multiwell plate with tags and

attached sensors



Sensor tags



Overall system

Sensors on tags within a

common rack

Tags with whole measurement

circuitry

Reader with connection to data

acquizition unit (e.g. PC)

Wireless energy and data

transfer



Conditions

Cell line 3T3 Clone A31 (Mouse

fibroblast)

Cell culture medium DMEM with

10% Fetal Calf Serum (FCS)

~ 300µl per sensor device

Measurement period of about 10

minutes



0 24 48 72 t [h]-24

3 Cell samples

Blank

1.0

0.5

1.5

2.0

2.5

3.0

Re

{}

[a.u

.]Z

sen

s

Measurements on 3T3 fibroblasts

50000 cells per sensor („>“ 100% confluence)

Blanks

Cells

Seeding

Medium

exchange



Measurements on hMSC

50000 cells per sensor, cell differentiation

1.5 3.0 t [d]1.0 2.0 2.50.50

-15

-5

0

Re{

} . 10

[m

]Z

sen

s

28

-10

0

D





3 4 7 t [d]2

0

10

15

5 61

5

0

Re{

} . 10

[m

]Z

sen

s

28

D

Measurements on hMSC

50000 cells per sensor, cell differentiation





There is urgent need for investigation cell wall characteristics by a quick method.


Acknowledgments

Collaborators Funding organizations

NAMDIATREAM project

(EU NMP4-LA-2010-246479)

SPONSORED BY THE

Project # 823985

mnt-era.net project MinoLab

http://selectiveantibodies.com/www

http://selectiveantibodies.com/www

http://www.tu-braunschweig.de/

http://www.tu-braunschweig.de/


www.ait.ac.at/profile/detail/Heer-Rudolf

[email protected]

Thanks for your attention.

Lab-on-a-chip...Funded by the European Union - GA 737089 Kick-Off Meeting, Barcelona, Jan 17-18, 2017 1 ChipScope project kick-off meeting Lab-on-a-chip Rudolf Heer, [email protected],

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