Microsenzorii stocastici ca noi

instrumente de masurare a

substantelor de importanta biologica

PN-II-ID-PCE-2011-3-0570

Raluca-Ioana van Staden

Laboratorul de Electrochimie si PATLAB Bucuresti

Institutul National de Cercetare pentru Electrochimie si Materie Condensata

PATLAB

Obiective PATLAB

O1 Identificarea celor mai bune materiale nanostructurate pentru designul microsenzorilor stocastici; O2 Evaluarea diferitilor parametrii care pot afecta caracteristicilor de raspuns ale microsenzorilor stocastici cum ar fi, natura matricei, pH-ul si taria ionica, electrolitii; O3 Aplicarea noilor microsenzori stocastici la determinarea moleculelor de importanta biologica, cum ar fi hormoni tiroidieni, biomarkeri pentru obezitate, etc.; O4 Dezvoltarea de instrumente pentru scanarea substantelor de importanta biologica; O5 Dezvoltarea unei baze de date pentru analiza calitativa utilizand microsenzori stocastici; O6 Integrarea tinerilor cercetatori in echipa laboratorului.

Test de scanare pentru obezitate

PATLAB

Electroactive

material

Matrix Calibration

equation

Linear

conc.

range

toff

(s)

Sensitivity (mg/s . mL)

P DP 1/ton=0.02 + 1.96 x 102 x c

R = 0.9965 0.125pg/mL-0.125µg/mL 6.4 1.96x105

MD DP 1/ton=0.06 + 1.69 x 102 x c

R = 0.9966 1.25pg/mL-0.125µg/mL 3.9 1.69x105

MD Graphite

1/ton=0.04 + 1.30 x 102 x c

R = 0.9791 1.25pg/mL-0.125µg/mL 5.2 1.30x105

MD Graphene

1/ton=0.03 + 2.26 x 103 x c

R = 0.9993 0.125pg/mL-12.5pg/mL 6.1 2.26x106

MD CNT 1/ton=0.05 + 0.07 x 102 x c

R = 0.9091 12.5pg/mL-1.25µg/mL 3.9 7.00x103

MD C60

1/ton=0.02 + 1.21 x 102 x c

R = 0.9998 1.25pg/mL-0.125µg/mL 7.4 1.21x105

Leptin PATLAB

PAI-1

Electroactive

material

Matrix Calibration

equation

Linear

conc.

range

toff

(s)

Sensitivity (mg/s . mL)

P DP 1/ton=0.04 + 3.9 x c

R = 0.9626 5ng/mL – 5µg/mL 7.2 3.90x103

MD DP 1/ton=0.06 + 3.8 x c

R = 0.9966 50ng/mL – 5µg/mL 3.6 3.37x103

MD Graphite

1/ton=0.01 + 9.43 x 103 x c

R = 0.9998 50pg/mL – 5ng/mL 9.9 9.43x106

MD Graphene

1/ton=0.07 + 4.50 x 102 x c

R = 0.9745 50pg/mL – 50ng/mL 2.6 4.50x105

MD CNT 1/ton=0.05 + 3.31 x 102 x c

R = 0.9695

500pg/mL –

50ng/mL 2.8 3.31x105

MD C60

1/ton=0.02 + 1.2 x 101 x c

R = 0.9996 50ng/mL – 5µg/mL 5.6 1.20x104

PATLAB

Scanarea probelor de sange pentru leptin.

Sample Leptin, pg/mL

P/DP MD/DP -CD/DP MD/C60 MD/

Graphite

MD/

Graphene

MD/CNT

1 10.1 15.0 11.7 15.6 9.87 10.0 10.4

2 14.0 19.0 12.0 10.0 10.0 14.0 16.0

3 1.00 1.00 1.46 1.73 1.35 1.60 1.36

4 10.0 10.0 14.8 19.0 11.7 10.0 14.0

5 3.00 3.00 3.49 2.80 3.30 3.73 3.30

PATLAB

Scanarea probelor de sange pentru PAI-1.

Sample PAI-1, pg/mL

P/DP MD/DP -CD/DP MD/C60 MD/

Graphite

MD/

Graphene

MD/CNT

1 108 119 99.0 106 107 105 109

2 105 109 97.0 103 136 105 100

3 146 180 130 120 135 100 136

4 466 441 410 440 488 420 490

5 508 500 538 573 563 510 560

PATLAB

Este necesara o metoda de scanare a salivei

copiilor pentru BPA.

*http://wakeup-world.com/2014/05/11/10-things-to-know-about-bpa-bisphenol-a/ *http://www.stats.org/stories/2009/science_suppressed_BPA_intro_jun12_09.html

*

BPA poate contribui la obezitatea copiilor

PATLAB

Caracteristicile de raspuns ale microsenzorilor stocastici

Matricea Materialul

Electroactiv

Ecuatia de calibrare si

coeficientul de corelatie*

Domeniul

linear de

concentratie

(mol/L)

toff (s) Sensibilitatea

(s mol L-1)

Graphene

MD

1/ton= 0.04 + 3.62 x 1011 x C

r=0.9999 10-15 – 10-13 6.6 3.62 x 1011

CNT 1/ton= 0.10 + 6.37 x 1011 x C

r=0.9999 10-15 – 10-13 8.2 6.37 x 1011

C60 1/ton= 0.04 + 1.33 x 1010 x C

r=0.9994 10-14 – 10-12 9.2 1.33 x 1010

PATLAB

Microsenzor

bazat pe

1 2 3 4 5 6 7 8 9 10 11 12

MD/C60 0.11 0.52 0.86 0.38 0.73 0.40 0.01 0.27 0.22 0.17 0.02 0.01

MD/CNT 0.14 0.53 0.87 0.39 0.74 0.46 0.01 0.26 0.23 0.15 0.01 0.03

MD/Graphene 0.15 0.56 0.80 0.36 0.85 0.56 0.01 0.20 0.18 0.20 0.03 0.02

ELISAa 0.12 0.56 0.87 0.45 0.74 0.56 -b 0.25 0.34 0.43 -b -b

Determinarea BPA (µg/mL) in saliva

copiilor

PATLAB

time (sec)

0 200 400 600 800 1000 1200 1400

I (A

)

-400x10-12

-300x10-12

-200x10-12

-100x10-12

0

100x10-12

200x10-12

300x10-12

400x10-12

toff

ton

Time (sec)

0 200 400 600 800 1000 1200 1400

I (A

)

-300x10-12

-200x10-12

-100x10-12

0

100x10-12

200x10-12

300x10-12

400x10-12 toff

ton

time (sec)

0 200 400 600 800 1000 1200 1400

I (A

)

-150x10-12

-100x10-12

-50x10-12

0

50x10-12

100x10-12

150x10-12

ton

toff

MD/C60 MD/GRAPHENE

MD/CNT

Diagrame obtinute pentru determinarea BPA in saliva copiilor

PATLAB

Test de scanare pentru DHT, E2, T2

Caracteristicile de raspuns ale microsenzorilor stocastici

Microsenzori bazati pe DP

si

Ecuatiile de calibrare si

coeficientii de corelatie (r)

Domeniul linear de

concentratie

(mol/L)

toff

(s)

Sensibilitatea

(s/mol L-1)

DHT

P 1/ton=0.05 + 1.86 x 1010 x c

r=0.9853 10-14 – 10-12 4.0 1.86 x 1010

MD 1/ton=0.08 + 2.65 x 1011 x c

r=0.9996 10-15 – 10-13 1.8 2.65 x 1011

-CD 1/ton=0.05 + 6.92 x 108 x c

r=0.9825 10-13 – 10-11 5.1 6.92 x 108

T2

P 1/ton=0.065 + 1.27 x 106 x c

r=0.9942 10-10 – 10-8 5.3 1.27 x 106

MD 1/ton=0.061 + 1.95 x 108 x c

r=0.9989 10-12 – 10-10 3.5 1.95 x 108

-CD 1/ton=0.069 + 1.27 x 107 x c

r=0.9999 10-11 – 10-9 3.1 1.27 x 107

E2

P 1/ton=0.03 + 2.51 x 109 x c

r=0.9987 6.59 x 10-14 – 6.59 x 10-12 7.4 2.51 x 109

MD 1/ton=0.02 + 3.50 x 105 x c

r=0.9966 6.59 x 10-11 – 6.59 x 10-8 8.5 3.50 x 105

-CD 1/ton=0.06 + 1.83 x 109 x c

r=0.9976 6.59 x 10-14 – 6.59 x 10-12 5.2 1.83 x 109

Determinarea hormonilor (pg/mL) in saliva

copiilor

aELISA (metoda standard). bELISA nu a putut determina analitul.

Microsenzor

bazat pe DP

si

1 2 3 4 5 6 7 8 9 10 11 12

F F F F F F M M M M M M

T2

-CD/DP 3.1 0.3 1.2 1.3 3.3 1.7 2.3 2.1 1.0 1.1 1.7 1.1

P/DP 3.3 0.3 2.2 1.7 3.9 1.9 2.7 1.9 1.3 1.4 1.7 1.3

MD/DP 3.2 0.3 1.9 1.1 3.3 1.1 2.2 1.8 1.1 1.2 1.6 1.5

ELISAa -b 0.7 1.1 0.4 0.6 0.9 1.0 1.4 1.1 0.5 0.6 0.3

DHT

-CD/DP 0.3 0.3 0.3 0.2 0.1 0.3 0.3 0.3 0.3 0.1 0.2 0.2

P/DP 0.2 0.2 0.4 0.2 0.1 0.4 0.2 0.2 0.2 0.2 0.2 0.1

MD/DP 0.2 0.3 0.3 0.1 0.1 0.4 0.3 0.3 0.2 0.1 0.1 0.3

ELISAa -b -b 0.2 0.4 0.2 0.5 0.4 0.7 0.8 0.4 1.1 0.3

E2

-CD/DP 1.9 2.9 1.1 1.1 2.3 2.2 1.8 1.7 0.2 1.0 0.8 0.9

P/DP 1.7 2.3 1.0 1.0 2.2 2.0 1.4 1.5 0.6 1.3 1.0 1.0

MD/DP 1.1 2.7 1.2 1.5 2.6 1.5 1.6 1.4 0.7 1.0 0.8 1.6

ELISAa 1.8 1.7 1.3 1.1 1.5 2.0 1.3 1.7 0.5 1.1 1.0 0.9

Analiza DHT, E2, T2 in saliva utilizand microsenzorul bazat pe P/DP

Analiza DHT, E2, T2 in saliva utilizand microsenzorul bazat pe MD/DP

DHT T2 E2

Analiza DHT, E2, T2 in saliva utilizand microsenzorul bazat pe -CD/DP

HEPATITA B

Scanarea sangelui pentru

hepatita B si cancer !!!

PATLAB

Biomarkeri

• Cancer de san:

- CA15-3

• Hepatita B:

- HBvAg

- Acid D-pipecolic (D-PA)

Caracteristicile de raspuns ale senzorului

multimode bazat pe MD/DP, determinate in

modul stocastic.

Biomarker toff

(s)

Domeniul linear de

concentratie

Sensibilitate Ecuatie de calibrared

CA15-3 3.3 5 – 500 a 44mU s/mL 1/ton= 20.11 + 0.044xC;a

R= 0.9608

HBVAg 1.9 10-10-10-8 b 3.9 x 105 g s/mL 1/ton= 0.05 + 3.9x105xC;b

R= 0.9997

D-PA 4.9 10-12-10-10 c 3.0 x 108 mol s/L 1/ton= 0.02 + 3.0x108xC;c

R= 0.9488

aU/mL; bg/mL; cmol/L; d<ton> = s.

Caracteristicile de raspuns ale senzorului

multimode bazat pe MD/DP, determinate in

modul DPV.

Biomarker E

(mV)

Linear

concentration

range

Sensitivity Equation of calibration

CA15-3 780 5 – 500 a 5pA U/mL H= 0.3 + 5.0xC;a,d

R= 0.9382

HBVAg 526 10-12- 5x10-9 b 2.9nA g/mL H= 3.1 + 2.9xC;b,e

R= 0.9976

D-PA 350 10-11-10-8 c 155.4pA mol/L H= 0.2 + 0.16xC;c,e

R= 0.9915

aU/mL; bg/mL; cmol/L; d<H>=pA; e<H>=nA

E(mV)

0.68 0.70 0.72 0.74 0.76 0.78 0.80 0.82 0.84 0.86 0.88

I(A

)

-5.0e-11

0.0

5.0e-11

1.0e-10

1.5e-10

2.0e-10

2.5e-10

3.0e-10

3.5e-10

E(mV)

0.25 0.30 0.35 0.40 0.45 0.50 0.55 0.60 0.65

I(A

)

-2.0e-10

0.0

2.0e-10

4.0e-10

6.0e-10

8.0e-10

1.0e-9

1.2e-9

E(mV)

0.1 0.2 0.3 0.4 0.5 0.6

I(A

)

0.0

2.0e-10

4.0e-10

6.0e-10

8.0e-10

1.0e-9

1.2e-9

1.4e-9

1.6e-9

1.8e-9

D-PA 1x10-12mol/L

Analiza calitativa a biomarkerilor in probele de

sange.

Biomarker Sample 1 Sample 2 Sample 3

Stochastic

mode

Standard

method

Stochastic

mode

Standard

method

Stochastic

mode

Standard

method

CA15-3 + + + + + +

HBVAg + + + + + +

D-PA + + + + + +

E(mV)

0.0 0.2 0.4 0.6 0.8 1.0

I(A

)

-2e-10

0

2e-10

4e-10

6e-10

8e-10

1e-9

Determinarea cantitativa a biomarkerilor direct

din sange.

Biomarker Sample 1 Sample 2 Sample 3

M1 M2 Std M1 M2 Std M1 M2 Std

CA15-3a 10.2 10.8 9.6 43.7 42.9 42.0 87.2 87.9 86.0

HBVAgb 420 419 417 327 327 324 529 528 524

D-PAc 325 326 320 219 216 211 625 627 620

CA15-3 HBVAg

D-PA

Sample 2

t(s)

0 100 200 300 400

I(A

)

-1e-10

-8e-11

-6e-11

-4e-11

-2e-11

0

2e-11

4e-11

6e-11

8e-11

1e-10 D-PA HBVAg CA15-3

Diseminarea rezultatelor

Capitol in carte:

Mechanism of potential development for potentiometric sensors, based on modeling of interaction between electrochemically active compounds from the membrane and analyte

R.I. Stefan-van Staden

Chapter 4, pp.131-154 in

Chemical Sensors Simulation and Modeling

Volume 5: Electrochemical Sensors (Ghenadii Korotcenkov, Ed)

Published in Sensors Technology Series (Joe Watson, Ed.)

Momentum Press, LLC, New York, 2013.

Diseminarea rezultatelor 1. Stochastic sensors based on nanostructured materials used in the screening of

whole blood for hepatitis B. R.I. Stefan-van Staden, I. Moldoveanu J Electrochem Soc, 161(2), B3001-B3005, 2014

2. New tool for screening of whole blood for early detection of breast cancer antigen (CA153). R.I. Stefan-van Staden, J.F. van Staden J. Mod.Med.Chem., 1(2), 86-91, 2013

3. Enantioselective Surface Plasmon Resonance Sensor Based on C60 Fullerene-Glutathione Self-Assembled Monolayer (SAM). R.I. Stefan-van Staden. Chirality, 26(3), 129-131, 2014

4. New Multimode Sensors based on Nanostructured Materials for Simultaneous Screening of Biological Fluids for Specific Breast Cancer and Hepatitis B Biomarkers. R.I. Stefan-van Staden, I. Moldoveanu J Electrochem Soc, 161(4), B45-B48, 2014

5. A new genetic test for obesity, based on stochastic sensors. Raluca-Ioana Stefan-

van Staden, Livia Alexandra Gugoasa. J Electrochem Soc, 161(9), B167-B170, 2014

6. Screening of children saliva samples for bisphenol A using stochastic, amperometric and multimode microsensors. R.I. Stefan-van Staden, L.A. Gugoasa, B. Calenic, J.F. van Staden, J Legler Analytical Chemistry Research, 1, 000, 2014

Diseminarea rezultatelor

1. Analysis of L-thyroxine and 3,3’,5-triiodo-L-thyronine using potentiometric

microsensors. Iuliana Moldoveanu, Raluca-Ioana Stefan-van Staden, Jacobus

Frederick van Staden, Gabriel Lucian Radu U.P.B. Sci. Bull., In Press.

2. Amperometric microsensors based on inulins for the assay of L-T3 and L-T4.

Livia Alexandra Gugoasa, Raluca-Ioana van Staden, Grzegorz Bazylak,

Jacobus Frederick van Staden, Gabriel-Lucian Radu U.P.B. Sci. Bull., In Press.

Implicarea tinerilor

• Iuliana Moldoveanu – doctorand

• Livia Alexandra Gugoasa – doctorand

• Ionela Raluca Comnea – doctorand

• Ramona Georgescu - doctorand

• Dr Bogdan Calenic – postdoc

• Oana Elena Stoica – asistent tehnician

• Anita Girbea – masterand

• Radu Constantin Iacomin - masterand