neso sojic - bordeaux.udppc.asso.fr

Neso Sojic [email protected]

«Un peu de soleil dans la mer ou comment des réactions

redox conduisent à la détection ultrasensible »

ECL…..diagnostics…

D. M. Rissin et al., Nat. Biotech., 2010, 28, 595

● Cancer: a 1-mm3 tumor composed of a million cells that each secrete

5,000 proteins into 5 liters of circulating blood translates to a

concentration of ~2 × 10−15 M (or 2 fM).

● Infection: serum from individuals recently infected with HIV contains

10–3,000 virions per ml, resulting in estimated concentrations of the

p24 capsid antigen ranging from 50 × 10−18 M (50 aM) to 15 × 10−15 M

(15 fM).

Detection of protein biomarkers ► to differentiate between healthy

and disease states, and to monitor disease progression.

𝒄𝒐𝒏𝒄𝒆𝒏𝒕𝒓𝒂𝒕𝒊𝒐𝒏 = 𝒂𝒎𝒐𝒖𝒏𝒕

𝒗𝒐𝒍𝒖𝒎𝒆

Global Immunoassays Market:

by applications: by technology:

Oncology

Infectious Diseases

Inflammatory Diseases

Cardiology

Neurology

Bone and Mineral

Endocrinology

Autoimmunity

Toxicology

Hematology (Anemia)

Neonatal Screening Neurology

…

Electrogenerated ChemiLuminescence (ECL)

Immunoassays Commercialised for clinical applications:

Immunology

Toxicology

Metabolic Application

Inflammation

Alzheimer’s Disease

Meso Scale Discovery, www.mesoscale.com

Roche Diagnostic Corp., www.roche.com 4

Roche commercializes more than 130 immunoassay tests for cardiac and

infectious diseases, maternal care, thyroid, and tumor markers, etc.

1,300,000,000 Elecsys tests based on ECL

technology sold in 2013. Source: Roche

Diagnostics-Business Overview 2014 Report

http://www.mesoscale.com/

http://www.roche.com/

July 25th, 2003

Roche Buys Igen in $1.4 Billion Deal; Creates New Company

Roche has settled its patent dispute with Igen by agreeing to buy the company for $1.4 billion and

creating a new subsidiary. The deal gives Roche full rights to Igen's electrochemiluminescence (ECL)

technology, which had been the object of contention between the companies.

According to Roche, it will pay Igen shareholders $47.25 in cash and one share of the newly formed

company for each share of the old Igen stock. The new company will hold Igen's patents and operations

in life sciences, clinical testing and other industries. Roche will receive non-exclusive, perpetual rights

to use ECL in the in-vitro diagnostics market and sell ECL-based immunochemistry to point-of-

care and doctors' offices. Additionally, Roche will retain improvements it makes to ECL technology.

The deal settles a long-running dispute that culminated in a court decision handed about two weeks ago.

In it, the U.S. Court of Appeals had struck down a $486 million ruling against Roche but affirmed Igen's

right to terminate a licensing agreement to Roche. The litigation was inherited by Roche when it

acquired Boehringer Mannheim in 1998, which had an existing agreement with Igen. By settling with

Igen, Roche's diagnostic business, the largest in the world, assured that it would continue to have access

to the ECL technology, which accounted for $404 million of Roche's clinical diagnostic sales.

•Photoluminescence, a result of absorption of photons

•Fluorescence

•Phosphorescence

•Bioluminescence, emission by a living organism

•Chemiluminescence, a result of a chemical reaction

•Electrochemiluminescence (or electrogenerated chemiluminescence), a result of an

electrochemical reaction

•Crystalloluminescence, produced during crystallization

•Electroluminescence, radiative recombination of electrons and holes in a material, usually a

semiconductor

•Cathodoluminescence, a result of being struck by an electron

•Radioluminescence, a result of bombardment by ionizing radiation

•Sonoluminescence, a result of imploding bubbles in a liquid when excited by sound

•Thermoluminescence, the re-emission of absorbed light when a substance is heated

•Mechanoluminescence, a result of a mechanical action on a solid

•Triboluminescence, generated when bonds in a material are broken when that material

is scratched, crushed, or rubbed

•Fractoluminescence, generated when bonds in certain crystals are broken by fractures

•Piezoluminescence, produced by the action of pressure on certain solids

Different types of luminescence:

Internal

conversion

1) Chemiluminescence Resulting from Electrochemically Generated Species

David M. Hercules

Science 1964, 145, 808.

Chemiluminescence is reported from chemical species produced during electrolysis of aromatic

hydrocarbons in deoxygenated nonaqueous solvents. Emission occurs in the solution near the

cathode, and the emitting species is the excited singlet state of the hydrocarbon. Oxygen exerts a

severe quenching effect on the chemiluminescence. Possible mechanisms are discussed.

2) Electroluminescence in Solutions of Aromatic Hydrocarbons

Visco, R. E.; Chandross, E. A.

J. Am. Chem. Soc. 1964, 86, 5350.

3) Chemiluminescence of Electrogenerated 9,10-Diphenylanthracene Anion Radical

Santhanam, K. S. V.; Bard, A. J.

J. Am. Chem. Soc. 1965, 87, 139.

The origins of ECL……electrochemistry of aromatic hydrocarbons in organic solvents

L’oxydo-réduction……un grand “mystère”…

réducteur

oxydant

réduction

oxydation

anode

cathode

intensité

courant

potentiel

tension

Faraday

pile

batterie

électrolyse

Ox + e- → Red

forme oxydée

forme réduite

Ox1 + Red2 → Red1 + Ox2

Marcus model for the electron-transfer reaction

Rudolph A. Marcus. Nobel prize 1992

Rudolph A. Marcus developed an original theory to explain the rate of electron-transfer

reactions, linking the thermodynamics of the process to its kinetics.

His original article (1956) referred to the self exchange

reaction in aqueous solution :

Fe2+ + Fe3+ → Fe3+ + Fe2+

where the reorganization energy is solely due to

reorganization of water dipoles around solvated ions.

ELECTRON TRANSFER REACTIONS IN

CHEMISTRY: THEORY AND EXPERIMENT

R. A. Marcus - Nobel Lecture, December 8, 1992

Marcus model for the electron-transfer reaction

R. A. Marcus - Nobel Lecture, December 8, 1992

Electron transfer processes occur

over a very short time scale, thus

requiring rapid dissipation of a

large amount of energy into the

vibrational modes of the molecular

frameworks. This is rather difficult

for the reacting system, leading to

kinetic inhibition of direct formation

of the stable ground state products

A and D.

The formation of the excited states (A* or D*) is less exergonic and

correspondingly less thermodynamically favored, but the process may

be kinetically preferred because a relatively small amount of energy

needs to be vibrationally dissipated.

ECL can be understood in terms of competition of (at least) two

electron transfer reactions.

where A and D could be the same species, e.g., an aromatic hydrocarbon

S-ROUTE : « energy sufficient »system

Energy Requirements

The energy of the electron transfer reaction governs which excited states are

produced. Because the excitation energy is fundamentally closest to a

thermodynamic internal energy, the usual energy criterion for production of an

excited singlet state of energy ES (in electronvolts) is:

Frequently the criterion is given based on CV peak potentials (in volts) at T 298 K as:

Such reactions are sometimes called “energy sufficient,” and the reaction is said to

follow the S route.

Electrochemistry and ECL of aromatic molecules in organic solvents

Analytical signal: light intensity Cyclic amplification of the signal The intensity of the light emitted correlates with the concentration of Ru(bpy)3

2+ and of the co-reactant (TPA). Extremely sensitive detection technique.

Simplified mechanism: LIGHT (λ=615 nm)

The discovery of ECL in water with sacrificial co-reactant species

Mechanism?

Behave differently depending on concentration, pH, electrode materials,…

W. Miao, J.-P. Choi, A. J. Bard J. Am. Chem. Soc. 2002, 124, 14478

M. Zhou, J. Heinze,K. Borgwarth, C. P. Grover ChemPhysChem 2003, 4, 1241

R. M. Wightman, S. P. Forry, R. Maus, D. Badocco, P. Pastore J. Phys. Chem. B 2004, 108, 19119

A) B)

C)

Competitive mechanistic pathways leading to ECL

Diagnostics…………ECL

Immunoassays are an integral part of the healthcare practice. It is a

highly specific in vitro biochemical test that uses antigen-antibody

reactions to detect the concentration of different biomarkers.

Current immunoassays typically measure proteins at concentrations

above 10−12 M.

Sandwich format:

: label (enzyme, fluorophore,

ECL-active, radioisotope, etc.)

: biomarker (antigen)

Ru-TAGTM labels, beads, antibodies, and analytes are mixed in solution

and driven into the flow cell by simple fluidic path flow.

The flow cell is at the heart of the 2nd generation of Igen technology:

The complex molecules (made of Ru-TAG labels, antibodies, analyte, and

biotin) are captured by the magnetic beads coated with streptavidin.

Igen technology:

The magnetic field is applied to bring the beads closer to the electrode.

Igen technology:

When the electrical potential is applied to the electrode, ECL takes place.

The amount of light which depends on the analyte amount is measured by

the light detector.

Igen technology:

After measurement, the magnetic field is turned off and the beads are

released.

Igen technology:

After the beads are evacuated, the flow cell is washed and ready for

another measurement.

Igen technology:

No multiplexing (one analyte per test)

Time line of ECL

W. Miao Chem. Rev., 2008, 108, 2506

1964–1965: First experiments

1965: Theory

1966: Transients

1969: Magnetic field effects

1972: Ru(bpy)32+

1977: Oxalate

1981: Aqueous

1982: Ru(bpy)32+ polymer

1984: Ru(bpy)32+ label

1987: Tri-n-propylamine (TPrA)

1989: Bioassay

1993: Ultramicroelectrodes

2002: Semiconductive nanocrystals

2014: 3D ECL?

1 – Multiplexed bead-based ECL immunoassays

2 – Mechanism of bead-based ECL immunoassays

3 – ECL immunosensing at nanoelectrode arrays

4 – Stimuli-responsive ECL microgels

5 – Light-emitting swimmers and 3D ECL

sample

antigen

biotinylated

detection Ab

SA-Ru

complex

TPA

redox 3 mm

bead

Ru(bpy)3+2

gold electrode

antibody

antigen

biotin

streptavidin

The principle: ECL imaging as readout mechanism

Deiss, F.; LaFratta, C. N.; Symer, M.; Blicharz, T. M.; Sojic, N.; Walt, D. R. J. Am. Chem. Soc. 2009, 131, 6088

I. Multiplexed ECL Sandwich Immunoassays

Illumina technology: Multiplexed Fluorescence

Capture Antibody

Detection Antibody

Antigen

Biotin h

h

Microsphere

Streptavidin-fluorophore

Cytokine (or DNA)

Capture Microsphere

Random distribution

www.illumina.com

The decoding and the detection steps are based on fluorescence

Fluorescent label

Optical fiber bundle: Chemical etching:

Microstructured gold-coated electrode comprising 50 000 microwells


3 Antigens are detected: - IL8

- TIMP-1

- VEGF

Beads encoded with Eu3+

Antibodies specific for interleukin-8 (IL-8), tissue inhibitors of metalloproteinase 1

(TIMP-1) and human vascular endothelial growth factor (VEGF).


Fluorescence Mapping and false colored

(before incubation with StreptAvidin-Ru(bpy)3 2+)

Specificity (1): 3 kinds of beads, 1 Ag, 3 dAb (3 mg/mL for each)

IL8 : 50 ng/mL

TIMP-1 : Ag absent VEGF : Ag absent

ECL imaging

• Well-separated ECL spots locations of individual anti-IL-8 labeled beads

• ECL imaging resolved at the single-bead level

• ECL mechanism


Cross reactivity

Ag Mw

(kDa)

C

(ng/mL)

C

(nmol/L)

IL8 8 50 6

VEGF 42 1000 24

TIMP-1 28 100 4

3 mg/mL for each dAb in cocktail

IL8

VEGF

TIMP-1

-0.2

0

0.2

0.4

0.6

0.8

1

IL 8 (50ng/mL)

VEGF (1μg/mL)

TIMP-1 (100ng/mL)

IL8

VEGF

TIMP-1


Triplex : 3 kinds of beads, 3 Ag, 3 dAb

• ECL imaging as readout mechanism for multiplex sandwich assay

Fluorescence ECL

IL8 : 50 ng/mL TIMP-1 : 1.5 μg/mL VEGF : 1.5 µg/mL

Deiss, F.; LaFratta, C. N.; Symer, M.; Blicharz, T. M.; Sojic, N.; Walt, D. R. J. Am. Chem. Soc. 2009, 131, 6088


N

N

N

NN

N

Ru2+

HN

O

CH3

interface: where electron-transfer reaction occurs

II. Why ECL bead-based immunoassays are sensitive ?

few µm

electron: few nm

ECL imaging resolved at the single bead level

N

N

N

NN

N

Ru2+

HN

O

CH3

a) b) c)

M. Sentic, M. Milutinovic, F. Kanoufi, D. Manojlovic, S. Arbault, N. Sojic, Chem. Sci., 2014, 5, 2568


Top view:


PL ECL a)

b)

0.7 V 0.83 V 0.9 V 1.1 V

PS beads:

Magnetic

beads:


►The bead acts as an efficient lens focusing the light at their center.

► It concentrates the analytical signal and contributes to the sensitivity.

SIDE VIEW ECL imaging resolved at the single bead level

PL a)

ECL

PL b)

0.7 V 0.9 V 1.1 V

ECL

z PS:

Mag.:


Diameter: 12 µm

►ECL-emitting region was confined very close to the electrode surface.

► It extends only 3–4 µm along the z-axis.


0.0005

0.001

0.0015

0.002

0.0025

0.003

0

1 10-5

2 10-5

3 10-5

4 10-5

0 0.5 1 1.5 2 2.5 3 3.5 4

TPrA

TPrA

E

ECL intensity

EC

L in

ten

sity

(a.u

.)Co

nce

ntr

atio

n (

M)

z (µm)

TPrA●+ TPrA● ECL


Concentration and ECL profiles:

►Diffusion of both radicals resulting from co-reactant oxidation.

► It extends only 3–4 µm along the z-axis.

W. Miao, J.-P. Choi, A. J. Bard J. Am. Chem. Soc. 2002, 124, 14478

0

1000

2000

3000

4000

5000

6000

-8 -4 0 4 8 12 16 20

EC

L in

ten

sity (

a.u

.)

z (µm)

►Deprotonation constant rate of TPrA●+: 2920 s-1 (half-life time: 0.24 ms)

► Maximum ECL intensity occurs in the micrometric region where

concentrations of TPrA●+ and TPrA● radicals are locally the highest.

► Only the luminophores located in the 3 µm region next to the electrode

contribute to the ECL signal optimal size of the beads for the bioassays.

► New co-reactant with adequate redox potentials and appropriate radicals

life-time to to excite Ru(bpy)3 2+-labels located far from the electrode.


Distribution of ECL intensity: Comparison of experimental and simulated ECL :

Application: diagnosis of Celiac Disease (CD)

• Chronic, auto-immune disorder damaging Villi

of small intestine

• Genetically determined

• Affects bout 1% of the world population

• Triggered by ingestion of Gluten foods

US Pharmacist 2012

III. ECL immunosensing at nanoelectrode arrays

Green and Celier N. Engl. J. Med. 2007, 357:17

Basis for serologic

tests

Diagnosis

Serological (Anti-tTG/AGA)

ELISA ECL?

Intestinal

Biopsy

• Transducer – PC* templated Au NEEs

• tTG (tissue TransGlutaminase) immobilized on the PC

• Anti-tTG captured by tTG (in the sample)

• Biot-secondary Ab reacted with Anti-tTG

*PC- track-etched polycarbonate membrane


Au nanoelectrode arrays:

Analyte (Anti-tTG) conc. vs ECL

• ECL recorded for different [anti-tTG]

• Calibration surve:

• linear from 1 ng/mL to 5 µg/mL

• with R2 of 0.9944

• Better reproducibility :

• minimal surface oxide formation when scan is reversed at < 1.0 V

•Tests on real samples



A) ECL signals of

representative serum

samples

B) Bar graph showing ECL

intensity vs serum samples

from 5 CD patients

50

IV. Stimuli-Responsive ECL Microgels

- Optical properties

- Porosity

- Water content

- Electrochemical properties

Modification of the physical properties:

Stimuli-responsive microgels: “smart” nanomaterials

51

Stimulus

OFF

ON

Swelling ratio depends on 3 parameters:

• Solvent-polymer interactions

• Cross-linking density

• Charge density

Microgels: 3-dimensional networks with the particle size of 100-1000 nm

IV. Responsive Microgels: Dynamic Systems

Phase transition is accompanied by a reversible, discontinuous volume change

in response to infinitesimal changes in environmental conditions (t°, pH,

composition, light, etc.).

Swollen state

Collapsed state

Example of poly(N-isopropylacrylamide) (pNIPAM)

Cross-linked pNIPAM

DT

T < VPTT

Hydrophilic

Swollen state

T > VPTT

Hydrophobic

Collapsed state

Hydrophobic

group

Hydrophilic

group

Linear pNIPAM

IV. Thermo-Responsive Hydrogels

IV. Synthesis of the ECL Microgels

N

N

H3C

N

NN

N

Ru

2+/3+

O

HN

R1

O

HN

HN

O

+ +

R2

x % y %

Radical

polymerization N

N

H3C

N

N

N

N

Ru

CH2 CH

O

NH

CH

H3C CH3

CH2 CH

O

NH

CH2

NH

O

CHCH2

CH2 CH

2+/3+

1-x-y x y

x

Sample R1 R2 x in feed

(mol. %)

y in feed

(mol. %)

y in

microgel

(mol. %)

VPTT

dH (nm)

in the swollen state

(PDI)

dH (nm)

in the collapsed

state

pNIPAM-1 Isopropyl H 1.6 1.3 0.59 33°C 130

(0.07) 65

F. Pinaud, L. Russo, S. Pinet, I. Gosse, V. Ravaine, N. Sojic. J. Am. Chem. Soc. 2013, 135, 5517

IV. ECL of the Thermo-Responsive Microgels

Cyclic voltammetry (a) and ECL signal (b)of pNIPAM microgels in PBS and 10

mM TPA at 25 °C (blue) and 37 °C (red). The Ru concentration is 50 µM.

-1.00E-05

0.00E+00

1.00E-05

2.00E-05

3.00E-05

4.00E-05

5.00E-05

0.2 0.4 0.6 0.8 1 1.2 1.4

-1000

0

1000

2000

3000

4000

5000

6000

7000

0.2 0.4 0.6 0.8 1 1.2 1.4

Current: ECL:

►Small increase of the oxidation waves after VPTT

►ECL emission at the oxidation potential of Ruthenium centers (1.1 V)

►ECL and PL spectra are identical same MLCT excited state

►Large enhancement of the ECL intensity in collapsed state


►Reversible enhancement of the ECL signal

0

1 104

2 104

3 104

4 104

5 104

6 104

7 104

0 1 2 3 4 5 6 7 8

EC

L in

ten

sity (

a.u

.)

Collapsed state ( > VPTT)

Swollen state ( < VPTT)


IV. Characterization of the Microgel Particles

56

dH (nm)

Sample R1 R2 % Ru

(mol%)

VPTT (oC)

< VPTT > VPTT

pNNPAM n-propyl H 1.74 20 160 100

pNIPAM-1 Isopropyl H 0.59 33 130 65

pNIPAM-2 Isopropyl H 1.99 33 150 80

pNIPAM / pNIPMAM

Isopropyl H/CH3 0.5 40 195 105

60

80

100

120

140

160

180

200

10 15 20 25 30 35 40 45 50

Temperature (°C)

Dia

me

ter

(nm

)


60

80

100

120

140

160

180

20010 15 20 25 30 35 40 45 50

Dia

me

ter

(nm

)

a)

0

20

40

60

80

100

10 15 20 25 30 35 40 45 50

No

rma

lize

d E

CL

sig

na

l

Temperature (°C)

b)

►Large enhancement up to 100 occurs at the VPTT

►ECL (x20-100) = E (x2) +C (?) +L(-10%)??

►VPTT hydrophlic/hydrophobic transition

distance between Ru centers is modulated



60

80

100

120

140

160

180

20010 15 20 25 30 35 40 45 50

Dia

me

ter

(nm

)

a)

0

20

40

60

80

100

10 15 20 25 30 35 40 45 50

No

rma

lize

d E

CL

sig

na

l

Temperature (°C)

b)

0

0.5

1

1.5

2

2.5

3

3.5

2 3 4 5 6 7 8 9 10

pNIPAM-1pNIPAM-2

NIPMAMNNPAM

No

rma

lized

EC

L in

ten

sity

(a

.u.)

Distance (nm)

►Electron-hopping in the microgels?

► Chemical step? Bimolecular reactions to generate the

excited ECL state?

►Large enhancement up to 100 occurs at the VPTT

►ECL (x20-100) = E (x2) +C (?) +L(-10%)??

►VPTT hydrophlic/hydrophobic transition

distance between Ru centers is modulated


59

• ECL-Resonance Energy Transfer

Donor Accepto

r

d >> 1-10 nm

No energy transfer

Spectral overlap

ECL

Donor

Accepto

r

d ≈ 1-10 nm

Energy transfer

dipole-dipole interaction

400 500 600 700 800

0,0

0,2

0,4

0,6

0,8

1,0

wavelenth (nm)

Donor:

Ru complex

Acceptor:

Cyanine 5

F. Pinaud, R. Millereux, P. Vialar-Trarieux, B. Catargi, S. Pinet, I. Gosse, N. Sojic,V. Ravaine

J. Phys. Chem. B 2015, DOI: 10.1021/acs.jpcb.5b06920

• FRET ECL-RET

10 15 20 25 30 35 40 45 50 55

160

180

200

220

240

260

280

300

320

PL

In

ten

situ

(A

.U.)

Temperature (°C)

lex=646 nm lem=670 nm lem=670 nm lex=454 nm

Energy transfer

600 650 700 750

0

10

20

30

40

50

60

70

80

90

100

110

120

130

PL

In

ten

sity (

A.U

.)

Wavelength (nm)

15°C

40°C

15°C

40°C

660 670 680 6900

200

400

600

PL

In

ten

sity (

A.U

.)

Wavelength (nm)

15°C

40°C

15 20 25 30 35 40 45 50 5550

60

70

80

90

100

110

120

130

PL

In

ten

sity (

a.u

.)

Temperature (°C)

V. Light-Emitting Swimmers

(1966)

« Swimmers » – Controlled dynamic system

Design of micro- and nano-objets depends on the strategy chosen to drive the motion.

Chemical fueling: based on the use of a chemical fuel (H2O2)

Conical microengine

Gao W.; Sattayasamitsathis S.; Orozco J.; Wang J.; J. Am. Chem. Soc. 2011, 133, 11862


Principles of Bipolar Electrochemistry

(wireless method)


Loget G.; Zigah D.; Bouffier L.; Sojic N.; Kuhn A., Acc. Chem. Res. 2013, 46, 2513

Fosdick S. E.; Knust K. N.; Scida K.; Crooks R. M., Angew. Chem. Int. Ed. 2013, 52, 2

Principles of Bipolar Electrochemistry

(wireless method)


L

d DVmax = E.d/ L

Polarization of conducting object, exposed to an electric field, combine in a synergetic

way two redox reactions on at the extremities of the object , if ΔVmax is important

enough. This phenomenon can be used for propulsion of micro- and nano-objects.

Maximum polarization:

Loget G.; Zigah D.; Bouffier L.; Sojic N.; Kuhn A., Acc. Chem. Res. 2013, 46, 2513

Fosdick S. E.; Knust K. N.; Scida K.; Crooks R. M., Angew. Chem. Int. Ed. 2013, 52, 2


Asymmetric light-emitting electrochemical swimmer

Scheme of the set-up used for levitation experiments.

Cyclic voltammogram (blue) and ECL signal (red) of 0.5

mM Ru(bpy)32+ in the presence of 100 mM TPrA with

100 mM phosphate buffer (pH=7.4) on a home-made

glassy carbon electrode. Scan rate: 100 mV/s.


Asymmetric light-emitting electrochemical swimmer

The synergetic reduction of H2O at the

cathodic pole (bottom of the bead) and

oxidation of the ECL reagents at the anodic

pole (top of the bead) induces simultaneous

motion and light emission of the glassy

carbon bead in a capillary.

M. Sentic, G. Loget, D. Manojlovic, A. Kuhn, N. Sojic, Angew. Chem. Int. Ed. 2012, 51, 11284

H2O(l) + e- → ½ H2(g) + OH-(l)


Bipolar levitation of an ECL-emitting bead


Bipolar levitation of an ECL-emitting glassy carbon bead

Series of optical images showing

a GC bead emitting ECL at

different times during its motion.

It is exposed to an external

electric field of 25.5 V cm-1.

The bead is placed in a U-shaped

cell, filled with 100 mM PBS

containing 0.5 mM Ru(bpy)32+,

100 mM TPrA and a few drops

of surfactant.

Inset: Plot showing the height evolution h as a function of time t.

M. Sentic, G. Loget, D. Manojlovic, A. Kuhn, N. Sojic, Angew. Chem. Int. Ed. 2012, 51, 11284

CATHODE (–)

ANODE (+)

δ–

δ+

ECL (425 nm)

H2O

H2O2

H2O2 O2 Luminol

3-Aminophthalate*

MO

TIO

N

Luminol ECL – blue emitting ’’swimmer’’

L. Bouffier, D. Zigah, C. Adam, M. Sentic , Z. Fattah, D. Manojlovic, A. Kuhn, N. Sojic, ChemElectroChem, 2014, 1, 95

Tuning the wavelenght of light emission - luminophore

the motion (horizontal & vertical)


1s 4s 7s

MOTION


Electrochemiluminescent swimmers for dynamic glucose sensing

M. Sentic, S. Arbault, B. Goudeau, D. Manojlovic, A. Kuhn, L. Bouffier, N. Sojic. Chem. Comm., 2014, 20, 10161


Switching-on of ECL during the swimmer motion

in a vertical glucose concentration gradient

L

d

VI. 3D ECL

ECL

(Left) Optical image showing 30-µm diameter GC beads immobilized in a gel inside a

6-mm long capillary. (Right) Bulk ECL generated by all the beads.

ECL emission by the carbon beads in a capillary.

VI. 3D ECL

(Left) White-light image showing a dispersion of CNTs in a 6-mm long capillary.

(Right) Bulk ECL generated by the CNTs.

VI. Dual-color 3D ECL: Ru(bpy)32+ / Luminol

O2

Ru(bpy)32+

δ+ δ-

Ru(bpy)32+ *

H2O

DBAE

O2

Luminol δ+ δ-

3-Aminophthalate*

H2O

H2O2

O2

+

(A)

P

1 mm

(C) (B)

white light in the dark

A. de Poulpiquet, B. Diez-Buitrago, M. Milutinovic, B. Goudeau, L. Bouffier, S. Arbault, A. Kuhn, N. Sojic

ChemElectroChem 2015, DOI: 10.1002/celc.201500402

76

VI. Bi-enzymatic 3D ECL detection

2 H+

H2

Glucose

Glucono-

lactone

NAD+

NADH

Ru(bpy)32+*

Ru(bpy)32+

(B)

δ+ δ-

GDH

Choline

Betaine

O2

H2O2

3-Aminophthalate*

Luminol

(C)

H2O2

H2O

δ+ δ-

(A)

Graphite feeder electrodes

Glass capillary (ø 6 mm)

δ+ δ- e-

oxidation reduction

Ch.Ox. 3D ECL produced by a bi-enzymatic system in a suspension of

carbon microbeads. Images of the system under white light

before application of the electric field and ECL image in the dark.

• ECL swimmers

• Control of the ECL shapes

• Multiplexed ECL immunoassays

• ECL imaging resolved at the single-bead level

• Decorrelation of the decoding (fluorescence) and detection (ECL) processes

• Stimuli-responsive ECL microgel nanoparticles

•Biosensing swimmers

•Dynamic ECL immunoassays

•ECL mechanism at the single-bead level

•ECL-RET microgels

Conclusions & Perspectives

M. SENTIC, G. LOGET,

M. MILUTINOVIC, C. ADAM, F. PINAUD,

A. KUHN, V. RAVAINE, L. BOUFFIER, D. ZIGAH, S. ARBAULT

D. MANOJLOVIC

H. B. HABTAMU, P. UGO

D. R. WALT

F. KANOUFI

COLLABORATIONS

neso sojic - bordeaux.udppc.asso.fr

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