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.
Alam, Principles of Nanobisoensors, 2013
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!
Alam, Principles of Nanobisoensors, 2013
Dimensionideglioggettidarilevare…Why nanobiosensors:
Biomolecules are small
Alam, Principles of Nanobisoensors, 2013 9
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
13 Alam, Principles of Nanobisoensors, 2013
Lab on a Chip technologies
14 Alam, Principles of Nanobisoensors, 2013
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.
Alam, Principles of Nanobisoensors, 2013 15
ElettronicaConformabile:superfici3Detecnologieflessibili
SommarioSecondaParte- Schemageneralediunsensorechimico- Tipidianaliti
DimensionieproprietàdeglianalitiWhy nanobiosensors:
Biomolecules are small
Alam, Principles of Nanobisoensors, 2013 9
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…
Alam, Principles of Nanobiosensors, 2013 6
Small biomolecules: Glucose
C6H12O6
Mass .. 180 g mol-1, Size ~ 1nm
DNA:unpòmenopiccolo…
Alam, Principles of Nanobiosensors, 2013 7
H OH
PO4 5C
Sugar N
Base
PO4
5C Sugar
Biopolymers: DNA (deoxyribonucleic acid)
Charge ~q, Mass ~300 Dalton
Proteine:ancoramenopiccole…
Alam, Principles of Nanobiosensors, 2013 8
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
Alam, Principles of Nanobiosensors, 2013 9
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
Alam, Principles of Nanobiosensors, 2013
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.
Alam, Principles of Nanobiosensors, 2013 12
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→∞ →
Alam, Principles of Nanobiosensors, 2013
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)
Alam, Principles of Nanobiosensors, 2013 4
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*
Alam, Principles of Nanobiosensors, 2013
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
Alam, Principles of Nanobiosensors, 2013
SensitivitàSensitivity are similar
7
~ln
s
s
NS
Nª º« »¬ ¼
~ s
s
NS
Nª º« »« »¬ ¼
~ln
s
s
NS
Nª º« »¬ ¼
Alam, Principles of Nanobiosensors, 2013
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
3 Alam, Principles of Nanobiosensors, 2013
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
Alam, Principles of Nanobiosensors, 2013 4
TempodirispostaSidimostracheesisteunarelazionegeneraletraN,numerodimolecolecatturatesullasuperficie,ρ0,concentrazionedell’analitainsoluzioneet,tempodatada:
Nanobiosensors are highly sensitive
Is there something fundamental about the geometry?
aM mM µM nM fM pM
3 Alam, Principles of Nanobiosensors, 2013
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
7 Alam, Principles of Nanobiosensors, 2013
Tempodirisposta:sensoreplanare
Operation of a planar sensor
8
( ) ?N t =
0,t ρ
Alam, Principles of Nanobiosensors, 2013
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=
Alam, Principles of Nanobiosensors, 2013
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)
Alam, Principles of Nanobiosensors, 2013
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
Alam, Principles of Nanobiosensors, 2013 12
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
Alam, Principles of Nanobiosensors, 2013
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
Alam, Principles of Nanobiosensors, 2013
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