I - Biosensori · Lab on a Chip technologies Alam, Principles of Nanobisoensors, 2013 14 ....

Biosensori-I

SlidestrattedaEDXonlinecourse“ElectronicBiosensors”byM.A.Alam

– PurdueUniversity

SommarioPrimaparte-  Sensoriesensitività-  Concentrazionicaratteristiche-  Dimensioniedensitàdeglianaliti-  Sensoriemicrotecnologia-  Applicazioni

ParametrimisurabiliSensors and Science

3

Infrared

Red

Violet

Ultra-violet

16Hz-

28kHz

Camphor

Musk

Flowers

Mint

Ether

Acrid

Putrid

Cold

Heat

Contact

Pain

Salt

Sour

Sweet

Bitter

Umami

Electron microscope

Radio telescope

Physical

Bio-chemical

Sensor

10k@30cm2

~3 mm2.

50e6@

2.5cm2

Alam, Principles of Nanobisoensors, 2013

Esempidisensori(osistemi)permisurebiomedicheBiosensors are everywhere

… but these are not nanobiosensors! Alam, Principles of Nanobisoensors, 2013 4

Qualesensibilitàèrichiestanellemisurebiomediche?

NIEHS-NIH website.

Why nano-biosensors: low concentrations

mM fM

Alam, Principles of Nanobisoensors, 2013 5

AnalisidelsangueAlam’s Recent Blood Test

• Sodium 139 mM/L • Potassium 4.3 mM/L • Chloride 103 mM/L • CO2 24 mM/L • Glucose 89 mg/dL • Creatinine 17 mg/dL • pH 7.5 • WBC 9.2 k/cumm • RBC 4.3 million/cumm

7

Most concentrations are in mM/L.


ConcentrazioniMicro, pico, femto Molar ?!

8

1M = 6x1023 molecules/liter ~ 1x1015/(100 um)3 box

1 fM ~ 1 1 pM ~ 1000 1 uM ~ 1 billion

Single grain of salt in several Olympic-sized swimming pools!


Dimensionideglioggettidarilevare…Why nanobiosensors:

Biomolecules are small


Water Antibody Bacteria A Period Glucose Virus Cancer Cell Tennis Ball

They have different mass, charge, and electron affinity

…edimensionideisensoriA short history of nanobiosensors

10

Glucose pH-meter

Vacuum tube

MOSFET IC

Protein/DNA Virus/bacteria PCR

Genome sequencers

10 Alam, Principles of Nanobisoensors, 2013

Obiettivo:MedicinaPersonalizzata

12

Why does it matter: Personalized medicine

A (Adenine) – T (Thymine) or C (Cytosine) – G (Guanine)

~3,200,000,000 (Human)

~4,600,000 (E. Coli)

~9,700 (HIV)

Length of Genome

chromosome

Human genome sequencing is indispensable in personalized medicine.

base pairs

Obiettivo:Integrazionecondispositivimobili

‘More than Moore’ Technologies: Integrating sensors with mobile devices


Lab on a Chip technologies


Conformal Electronics: Heart

D.-H. Kim et al., Materials for multifunctional balloon catheters with capabilities in cardiac electrophysiologial mapping and ablation therapy, Nature Materials, 10, p. 316, 2011.


ElettronicaConformabile:superfici3Detecnologieflessibili

SommarioSecondaParte-  Schemageneralediunsensorechimico-  Tipidianaliti

DimensionieproprietàdeglianalitiWhy nanobiosensors:

Biomolecules are small


Water Antibody Bacteria A Period Glucose Virus Cancer Cell Tennis Ball

They have different mass, charge, and electron affinity

BiomolecolediinteresseBiomolecules

• Chemical Indicators – Glucose (diabetes) – Nitric oxide (Parkinson)

• Elements of life – DNA (Genetic) – Protein (disorders)

• Invaders – Virus (infection) – Bacteria (infection) – DDT (disrupts cells function)

Alam, Principles of Nanobiosensors, 2013 4

Charges Mass Redox potential Optical index

Biomolecolediinteresse:moltopiccolo…


Small biomolecules: Glucose

C6H12O6

Mass .. 180 g mol-1, Size ~ 1nm

DNA:unpòmenopiccolo…


H OH

PO4 5C

Sugar N

Base

PO4

5C Sugar

Biopolymers: DNA (deoxyribonucleic acid)

Charge ~q, Mass ~300 Dalton

Proteine:ancoramenopiccole…


H OH

Biopolymers: Protein

Charge ..variable, Mass ~125 Dalton

Enzymes Hormones Tissue Transport molecules

C

H

N H

H C

O

OH

Amino Carboxyl R

ProteinecomebiomarcatoriProtein Biomarkers for Cancer

• PSA for prostrate cancer

• Cardiac Troponin T (CTnT) for heart attack

• Phosphorylation of histone protein (Y-HYAX)

for ionizing exposure

• BRCA1 BRCA2 for breast & ovarian cancers


Virus:semprepiùgrandi… Viruses

10

No charges, but mass and tagging help identify them

Alam, Principles of Nanobiosensors, 2013

~100 nm

Batteri:ancorapiùgrandi… Bacteria

11

1 mm3 have tens of millions of bacteria 1 lb of our weight comes from bacteria


Calcolodiconcentrazioninelsangue:esempiodelfarmacocon

paracetamoloExample: How many molars in a 1000

mg Headache medicine?

• Tylenol is C8H9NO2. (C=12,H=1,N=14,O=16) • 151.16 gm contains 6.023x1023 molecules. • 1000 mg contains ~ 4x1021 molecules. • Blood volume is 5 liters. • Therefore, 8x1020 molecules/L, or the

concentration is 8e20/6e23=1.32 mM. • High concentration. Should work in minutes.


Quantoèimportanteconoscerelaconcentrazionediciòchesicercainuncertocampionebiologico?

Esempio:Unorganocolpitodacancrorilascianelsangue,aduncertostadiodisviluppo,10000cellule.Sonotanteopoche?Èpossibilerilevarle?10.000cellulesu5litri,significa2000celluleperlitro.Seestraggoncampionedisangueda1cm3,inmediaestraggo2cellule,seneestraggo1mm3,inmediaestraggo210^-3cellule.Questonumero(<1)rappresentainrealtàlaprobabilitàdipescare1cellula,praticamentenulla.

Quantoèprobabilecheilsensoreintercettil’analita?

S

T

M

1

2

3

14 Alam 2013 Alam, Principles of Nanobiosensors, 2013

Percorsodell’analitainsoluzione:randomwalkDiffusion Process – Why random walk

15 Alam, Principles of Nanobiosensors, 2013

2d Ddtρ ρ= ∇

Appendix: Derivation of the Diffusion Equation

18

iρ 1iρ +1iρ −

1 11 1 1 1( ) ( ) ( ) ( ) ( ) ( )2 2 2 2i i i i i i

tt t t t t t tρ ρ ρ ρ ρ ρ

τ − +

∆+ ∆ − = + − −

2D x τ≡ ∆

1 12

( ) ( ) ( ) ( ) 2 ( )2

i i i i it t t t t tD

t xρ ρ ρ ρ ρ− ++ ∆ − + −

=∆ ∆

22

2

d dD D

dt dxρ ρ ρ= = ∇

( )2xD

τ∆

≡

Distanzamediapercorsa

16

Diffusion Distance

2

( , 0) ( 0)

dD

dtx t x

ρ ρ

ρ δ

= ∇

= = =

~x Dt

2 4( , )4

x DtNx t e

Dtρ

π−=

22

( , )2

( , )

x x t dxx Dt

x t dx

ρ

ρ

∞

−∞∞

−∞

= =∫∫

SommarioTerzaParte-  Tipidisensori-  Dimensioniegeometriedeisensori

Schemageneralediunsensore

Sensor

T

M Introducing the sensor

T T

Mèl’eventualeelettrododiriferimento

Primoproblema:catturadell’analitaCapture of molecules on sensor surface

0( )F s Rd k kdtN N NN ρ= − −

( )(( )) 1 F s Rk k tssN eN t ρ− += −

0F s

ssF s R

kNk k

Nρρ

=+

, 0F sk ρ→∞ →


Secondoproblema:rivelazionedell’analitaBiomolecules

• Chemical Indicators – Glucose (diabetes) – Nitric oxide (Parkinson)

• Elements of life – DNA (Genetic) – Protein (disorders)

• Invaders – Virus (infection) – Bacteria (infection) – DDT (disrupts cells function)


Charges Mass Redox potential Optical index

LabelledsensorsNanoscale biosensors: Labeled approach

C

A

G

A

T

Q1

Capture Probe

C

A

G

A

T

Q1

T

T

A

C

G Q1* C

A

G

A

T

Q2 C

A

G

A

T

Q3 C

A

G

A

T

Q1 C

A

G

A

T

Q2 C

A

G

A

T

Q3

Imaging

Optical detection scheme

DNA Microarray, DNA chip

T

T

A

C

G Q1*


Label-freesensorsThree types of label-free sensors

6

Cantilever

Mass to frequency

Gate

Amperometric

Chemical to current

Ref. & Aux. Electrode

Potentiometric

Charge to current

Fluid Gate


SensitivitàSensitivity are similar

7

~ln

s

s

NS

Nª º« »¬ ¼

~ s

s

NS

Nª º« »« »¬ ¼

~ln

s

s

NS

Nª º« »¬ ¼


Analogy to camera … similar megapixels!

Tempodirisposta-  Considerazionigenerali-  Limitiintrinsecidovutialladiffusione-  Limitilegatiallageometriadellasuperficie-  Conclusioni

TempodirispostaIltempodirispostadiunsensoreèperdefinizionel’intervalloditempocheintercorretralavariazionedelparametrodamisurareel’istanteincuiilsegnaleregistratodalsensorevariacomeconseguenzadellavariazionedelparametrostesso.

1 uM 1 pM 1 fM

Time

Response

6

Response or settling time defined

( )0( ) Fg D

s s sN t t N tρ= ≡ = ×

TempodirispostaDalmomentocheilsensorecontienesempreunostratodirecettorichehannoilcompitodicatturarel’analita,taletempodipendedaunlatodalledinamichedicatturaedall’altrodal“tempodiavvicinamento”dell’analitaalrecettore

Nanobiosensors are highly sensitive

Is there something fundamental about the geometry?

aM mM µM nM fM pM


0( )F sd k NdNt

N= − ρ

2d Ddt

= ∇ρ ρ

W

Settling time for biosensors

The diffusion-capture problem is very challenging, especially for complex capture surfaces


TempodirispostaSidimostracheesisteunarelazionegeneraletraN,numerodimolecolecatturatesullasuperficie,ρ0,concentrazionedell’analitainsoluzioneet,tempodatada:

Nanobiosensors are highly sensitive

Is there something fundamental about the geometry?

aM mM µM nM fM pM


Dovegèunparametrodipendentedallageometriadellasuperficiedicattura

0( )F sd k NdNt

N= − ρ

2d Ddt

= ∇ρ ρ

W

( ) ( )0

Fg DN tt ρ= DF = 2 DF = 1 1 < DF < 2

Fractal geometry allows simple solution

0F

s

kρ

→∞→

5

Tempodirisposta1 uM 1 pM 1 fM

Time

Response

6

Response or settling time defined

( )0( ) Fg D

s s sN t t N tρ= ≡ = ×

TempodirispostaChièN?NèilnumerodimolecoleintrappolatesullasuperficiedelsensoreChièρ0?Èlaconcentrazionedell’analitainsoluzione.Larelazionepuòessererovesciataereinterpretata:

A fundamental relationship of biosensor

0

32FD

s sN tρ−

−= ×

Like the Heisenberg relationship, but for a sensor …

Minimum number of analyte (depends on transduction)

Settling (response) time

Fractal dimension

Limits of detection

D=1 1<D<2 D=2


Tempodirisposta:sensoreplanare

Operation of a planar sensor

8

( ) ?N t =

0,t ρ


Tempodirisposta:sensoreplanare

0( )F sd

k NdNt

N= − ρ

2dD

dt= ∇

ρ ρ

Exact solutions for a planar sensor

, 0F sk ρ→∞ =

9

( )0( , ) erf 2x t x Dtρ ρ=


Equazionedidiffusione+equazionechedescriveladinamicadiintrappolamento

Tempodirisposta:sensoreplanareExact solutions for a planar sensor

( )0( , ) erf 2x t x Dtρ ρ=

( ) 2

0

2erfx yx e dy

π−≡ ∫

Particles captured

[ ]0 00

4( ) ( , )N t x t dx Dtρ ρ ρπ

∞

= − =∫10

0 1 2 3 4 50

0.2

0.4

0.6

0.8

1

erf(x)erf(x/2)erf(x/3)erf(x/4)


Soluzioneesatta:

Distanzamediadidiffusione

11

x

The concept of the diffusion distance

2dD

dt= ∇

ρ ρ

~x Dt

~x Dt1 2 3t t t< <

AndamentodiρApproximate Solution in 1D: Diffusion Distance

Exact

04( )N t Dtρπ

= × ×

0 1 2 3 4 50

0.2

0.4

0.6

0.8

1

erf(x)erf(x/2)erf(x/3)erf(x/4)

Dt

01( ) ~2

N t Dtρ× ×

Approximate

0ρ


Èpossibiletrovareun’opportunaapprossimazioneperlasoluzione:

AndamentodiρAddiritturaapprossimareulteriormente,pertrovareunastimadelrisultato

Approximate Solution in 1D: Diffusion Distance

13

Dt

01( ) ~2

N t Dtρ× ×

Approximate

0ρ

0( ) ~N t Dtρ ×

Approximate

0ρDt


TempodirispostadelsensoreplanareNelcasodiunsensoreperfettamenteplanare,l’esponentegvale1/2

Response time of a planar sensor

2

20

1~ ssNtD ρ

DF=2

14

( )1/2R Dt�

0

0

( ) ~

~

N t A R A

Dt A

× × ×

× ×

ρ

ρ

( ) 2/1ttN ∝

Diffusion slowdown Alam, Principles of Nanobiosensors, 2013

TempodirispostadelsensorecilindricoResponse time of cylindrical sensor

NW

0

1~ ssN atD ρ

DF=1

15

( )0

0

2 20 0

2

( ) ~2

~

N t a R a

Dt

ρ

ρ

π π

π

× × −

×

( ) 1ttN ∝

Diffusion slowdown absent?!

~R Dt


Èpiùveloce!

Nonc’èunrealevantaggioausaregeometrieconD<1Nanodots sensor offer no significant additional advantage!

Planar

Nanowire Nanodot

100s

11

Hahm et al., 2004, Zheng et al., 2005, Li et al., 2005, Kuznesow et al., 2006, Gao et al., 2007, Stern et al., 2007

Performance limit of biosensors

InConclusione:Unadeiparametridiinteresseperunsensoreèiltempodirisposta.Nelcasodisensoriobiosensoriattiamisurarelaconcentrazionediuncertoanalitainsoluzione,iltempodirispostaèdeterminatodaduemeccanismi:-  l’intrappolamentodell’analitasullasuperficie-  Ladiffusionedell’analitadallasoluzioneallasuperficieQualechesiailmeccanismoditrasduzione,ilsecondomeccanismoèsemprepresente.EsisteunaleggegeneralecheregolaladipendenzadiNdat.Questaleggehalastessaformaqualunquesiailmeccanismoditrasduzione,cambiasoloilparametroginfunzionedellacaratteristichegeometrichedelsensore.Sensoricongeometriacilindricahannotempidirispostapiùbassidiquelliageometriaplanare

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