Semiconductor Biosensing Technology

Semiconductor Biosensing Technology

Loading
Loading Social Plug-ins...
Language: English
Save to myLibrary Download PDF
Go to Page # Page of 52

Description: Effects of Semiconductor Power Sector Election of the Charge Density of Potential Measurement, Protein, Reference Electrode, Field Effect-Based Biosensing Device, FET chip, Insulated-gate and Extended-gate Field Effect Transistors, Biosensing Device: Application and Science, Electrical Detection of Bio-Molecular Recognition Events Using Biotransistor.

 
Author: Lee Mi Sakata (Fellow) | Visits: 1708 | Page Views: 3333
Domain:  Medicine Category: Equipment Subcategory: BioMEMS 
Upload Date:
Link Back:
Short URL: http://www.wesrch.com/medical/pdfME14GWGS4HYXR
Loading
Loading...



px *        px *

* Default width and height in pixels. Change it to your required dimensions.

 
Contents:


sakata@biofet.t.u-tokyo.ac.jp
2010/10/28

Biosensing method
~ > ~

~
> ~

>

>

~
> ~ () >

~ > ~ >

2010/10/28

Biomolecules Cells







DNA





2010/10/28

Field Effect-Based Biosensing Device






10-200 nm
VG

DNA

+



Si3N4 SiO2
eee e- n-Si

e-

2010/10/28

FET chip

gate area

sensing spot

10mm

gate area
340mm

gate area

1mm

polymer cover

300mm

4 gate-FET
FET chip
Si3N4/SiO2 gate insulator

Channel layout

- Gate width

1 mm ~ 10 mm
Source Drain

- Gate length 1 mm ~ 2400 mm

L=5 mm W=2400 mm

16 gate FET chip
2010/10/28

Insulated-gate and Extended-gate field effect transistors
a. Insulated-gate FET

b. Extended-gate FET
Vg
+ + + +

Vg
+ + eeee+ ee+





A Id


ee-



Id

Si

ee-

ee-

A

Si

2010/10/28

Biosensing device : Application and Science


DNA





()
2010/10/28



Electrical Detection of Bio-Molecular Recognition Events Using Biotransistor
Field Effect-Based Biosensing Technology (Field Effect Transistor; FET)

Cell functional analysis
DNA analysis
SNP genotyping DNA sequencing
Sakata, T. & Miyahara, Y. Angew. Chem. Int. Edit. 45, 2225 (2006).

Drug screening Tissue engineering

Sakata, T. et al., Analytical Chemistry, 80 (2008), 1493.
Sakata, T. et al., Microelectronics Engineering, in press (2008).

Sakata, T. et al., IEEJ, 2 (2007), 295.

Hybridization Intercalation Extension reaction DNA-nanoparticle conjugate
Sakata, T. & Miyahara, Y. ChemBioChem 6, 703 (2005). Sakata, T. & Miyahara, Y. Biosensors & Bioelectronics 22, 1311 (2007). etc.

Field Effect-Based Biosensing Techonology

Cell movement Function of membrane protein Differentiation of stem cell Embryo activity Apoptosis Sialic acid
Sakata, T. et al., JJAP., 47 (2008), 368. etc.

Protein analysis
Immunoassay Drug screening
Ueda, H. et al. Nat. Biotechnol., 14, 1714 (1996). Sakata, T. et al. Analytical Chemistry (2009)

Open sandwich assay Amiroyde fiber

2010/10/28

Bodysensing (in vitro and in vivo)

QT ()



iPS




()
2010/10/28

Cell



2010/10/28

Field Effect-Based Biosensing Devices
� �


� Original Interface - � Physically-structured interface - � Chemically-synthesized interface - � Biologically-induced interface -


� (iPSC2C12) �


� �



� Insulated-gate FET Ion sensitive FET (ISFET) � Extended-gate FET - ITO()



- ()
- ()
2010/10/28

2010 Nobel Prizes MEDICINE
The Father of the Test Tube Baby MEDICINE Robert G. Edwards, the 2010 Nobel Laureate in Physiology or Medicine, battled societal and establishment resistance to his development of the in vitro fertilization procedure, which has so far led to the birth of around 4 million people.

2010/10/28

Assisted reproductive technology (ART)





2010/10/28

Ion charge monitoring of embryo activity
CO2 + H2O HCO3- + H+

Ag/AgCl



CO2




VG
Si3N4 SiO2 n-Si eeeee-

H+
e
eee-

+ H+ H
e- e- e

H+
ee-

H+
eeee-

ee-

ee-

Si3N4 SiO2 n-Si

e-

e-

e-

pH variation
OH OH OH OH OH OH OH OH OH OH

Si3N4 SiO2

OH2 OH OH2 OH2 OH OH2 OH OH2 OH2 OH2 e- e- e- e- e+ + + +

+

+

+

e- e- e- e- e-

n-Si

2010/10/28

Simultaneous analysis by microscopy and biotransistor






(1 ml) � �





2009/8/10

PDMS

(20 ml)
2010/10/28




2cell

4-8cell

16cell




, 1cell 2cell 4cell 8cell 16cell ( (




24 01 2438 1 3850 1 5060 2 6070 2

6880 3 7482 3 4 5



35 (01

38 1

68-80 3846 5162 2 85 3 113135 5,6 1 113-135 8

120192 6

72 1632 7 72 5,6

813

2010/10/28

Real-time observation of mouse embryo activity in vitro fertilization

1cell

PDMS(200mm)



2010/10/28











2010/10/28





2010/10/28

(iPS)
iPS (Induced pluripotent stem cells) : APS0001 iPS-MEF-Ng-20D-17
Okita, K. et al., Nature 448: 313-317 (2007).

( )
iPS

iPS



iPS (+LIF(Leukemia Inhibitory Factor))

iPS (+LIF+Noggin)

(+Noggin � LIF)


2010/10/28

iPS-FET
H. Sugimoto, T. Sakata (2010).

iPS
100 mm iPS

(h)

2010/10/28

Apoptosis � Programmed cell death
Electrical detection of apoptosis (Programmed cell death)
Volume reduction Fragmentation of cells

Apoptosis
Activation of caspase Chromatin concentration DNA fragmentation
Induction of apoptosis TNF receptor

Microscopy

K+ channel

Cl-channel H2O

K+

Y. Okada et all., 2001. Journal of Physiology, 532.1, 3-16.

Incubator

Real-time FET analyzer
2010/10/28

Simultaneous analysis by microscopy and biotransistor
JST CREST





� �



2010/10/28

Apoptosis � Programmed cell death
Hela cellsTRAIL, Fas ligand (ligand for apoptosis)

Detection of K+ release at 3~5 h

gate area

100mm

Apoptosis of Hela cells on the gate sensing area
2010/10/28

Real-time observation of apoptosis on the gate
Control THIK-1(-)

THIK-1(+)

Control: Non-treated Hela cells
THIK-1(+): Increase of K+ ion channel THIK-1(-): Decrease of K+ ion channel

by transfection of RNA
2010/4/2
Sakata, Sakamaki, Sugimoto, (2010) JST CREST

2010/10/28

Chemically synthesized interface for apoptosis detection
15 pH 7.4 10

Surface potential (mV)

5

0 -5
Non-modified FET Non-modified FET Crown ether-modified FET Crown ether-modified FET

-10

-15 -5 -4 -3 -2 -1 Murakami, Sakata, et al. (2009)

K+ concentration log(M)

/
2010/10/28

Transparent extended-gate transistor

Indium Tin Oxide (ITO)

2010/10/28

Real-time and label-free monitoring of transporter function
Injection
hOATP-C
(Human Organic Anion Transporting Peptide C)

Sakata, T. & Miyahara, Y. Analytical Chemistry, 80 (2008), 1493.

Transporter

cRNA

expression Transportocyte

Estrone-3-Sulfate (E3S; C18H21O5SNa; 372.4) no expression Control Oocyte
Uptake clearance (pmol/oocyte)
0.6 0.5 0.4 0.3 0.2 0.1 VG

E3S

Control OATP-C RI labeled E3S


Ref. electrode



0
0 30 60

500 mm

Incubation time (min)

90

120

Si3N4 SiO2

e- e - e - e - e -

2010/10/28

Real-time and label-free monitoring of transporter function

20

Sakata, T. & Miyahara, Y. Analytical Chemistry, 80 (2008), 1493.
Ref. electrode
Ref. electrode

Interfacial voltage (mV)

Interface voltage (mV)

Control
VG
Si3N4 SiO2 e - e - e - e - eSi

20 15

Reference
VG
Si3N4 SiO2 e- e- e- e- eSi

15 10 5 0 0 -5 20

10 5 0 0 -5 20 40

injection

injection

40

60

80

60

80

Time (min)
Ref. electrode

Time (min)

Ion channel cell membrane

transporter

20
Interface voltage (mV)

15 10 5 0 0 -5

Oocyte with transporter
injection

VG

Si3N4 SiO2 e - e- e - e - e Si

drug Si3N4 electron eee- e- e- e

e-

e- ee-

20

40

60

80

Time (min)


2010/10/28

hOATP-C transporter
Organic Anion Transporting Peptide-C (OATP-C)
Wild type Mutant type

SNPs
name Europe Reference (NM_006446) N130D (A388G) V174A (T521C) N130D /V174A African American Frequency (%) Saharan African Chinese Japanese

*1a A:0.61,G:0.39 A:0.28,G:0.72 A:0.19,G:0.81 (n=114) (n=23) (n=90) T:0.92,C:0.08 (n=24) Africa T:0.98,C:0.02 (n=23) A:0.20,G:0.80 A:0.36,G:0.64 (n=69) (n=44) Asia T:0.88,C:0.12 (n=24)

*1b

*5

*15

2010/10/28

Radioisotope signal for hOATP-C transporter
OATP-C*1aOATP-C*15 (,60)
1.0

(L/Oocyte/60min)

Uptake clearance


Control OATP-C*1a

RI
1.0

0.8 0.6 0.4 0.2 0.0 4

***
(L/Oocyte/60min) Uptake clearance
0.8 0.6 0.4 0.2 0.0


OATP-C*1a OATP-C*15

20

Estradiol 17-D-glucuronide (mol/L)

0.8
(L/Oocyte/60min) Uptake clearance


Control OATP-C*15

0.6 0.4

4

20

Estradiol 17-D-glucuronide (mol/L)

0.2 0.0 4

***

Control

Wild type

Mutant type

20

Estradiol 17-D-glucuronide (mol/L)


2010/10/28

hOATP-C transporter mutation
Sakata, T. & Miyahara, Y. Analytical Chemistry, 80 (2008), 1493.

~FET measurement
VG

Ref. electrode

Oocyte FET Control

Transportocyte FET Wild type

Transportocyte FET Mutant type

Si3N4 SiO2 e- e- e- e- eSi

Si3N4 SiO2 e- e- e- e- eSi

Si3N4 SiO2 e- e- e- e- eSi

10

Interface voltage (mV) Surface potential (mV)

injection
8 6 4 2 0 0 -2 20 40

Wild type (OATP-C*1a) Mutant type (OATP-C*15) Control

Estradiol 17b-D-glucuronide concentration: 20mM
Room Temperature

2

60

80

Time (min)


2010/10/28

DNA

2010/10/28

DNA (Deoxyribo Nucleic Acid)
1953 DNA

1990 NIH

2003



2010/10/28

The Genome of Jim
May 2007 (Newsweek)

2010/10/28

DNA
Field-effect-based Biosensor/Biochip
Detection of various DNA recognition events using genetic FET
Hybridization
Ref. electrode

Intercalation
Ref. electrode

Primer Extension
Ref. electrode

Charged Nanoparticle
Ref. electrode

Au
VG VG VG
intercalator

VG

Si3N4 SiO2 Si

e- e- e- e- e-

Si3N4 SiO2 Si

e- e- e- e- e-

Si3N4 SiO2 Si

e- e- e- e- e-

Si3N4 SiO2 Si

e- e- e- e- e-

Field-effect-based DNA molecular recognition SNP genotyping, DNA sequencing, etc.
1. T. Sakata et al., Analytical Chemistry, 80, 1493, (2008). 2. T. Sakata et al., Angew. Chem. Int. Ed. 45, 2225, (2006). 3. T. Sakata et al., ChemBioChem, 6, 703, (2005). 4. T. Sakata et al., Biosensors & Bioelectronics, 21, 827, (2005). etc. 2010/10/28

Immobilization of oligonucleotide probe
Silanization of Si3N4 insulated gate surface

Glutaraldehyde

3-Aminopropyltriethoxysilane Si/SiO2/Si3N4 Si/SiO2/Si3N4

Sodium Cyanoborohyderide

Reaction between amino-group and aldehyde-group

Si/SiO2/Si3N4

Sodium Cyanoborohyderide

Si/SiO2/Si3N4

Immobilization of amino-group labeled oligonucleotide

Block of free aldehyde surface with glycine 2010/10/28

Direct Transduction of Extension Reaction into Electrical Signal
Extension reaction
T. Sakata and Y. Miyahara, Biosensors & Bioelectronics 22, 1311 (2007).

DNA polymerase and dNTP

Change of threshold voltage, VT (mV)

25
introduction of DNA polymerase

Threshold Voltage (VT) Negative Charges (DNA Molecules) +VT(positive)

20 15 10 5 0 0 50 100 150 200 250

time (s)

Charge density change due to extension reaction on gate could be detected into electrical signal using genetic FET.
2010/10/28

DNA
T. Sakata et al., Angew. Chem. Int. Ed. 45, 2225, (2006).

Factor VII R353Q
TGCACGGG

Change of threshold voltage, DVT (mV)

VG

3'-GCCATCACC-5' (9 bases) 5'-ACGTGCCCCGGTAGTGG-3' (17 bases)
a
A

15CA 13 G 11 9 7
T

b

A

C

A

G

C T

T

C

A

C T A

c

A

G

T

C

C G T A G C

d

A T

A

G

T

C G T A

G C

G C

G C

Si3N4 SiO2 Si

e- e- e- e- e-

Si3N4 SiO2 Si

e- e- e- e- e-

GGG

Si3N4 SiO2 Si

e- e- e- e- e-

Si3N4 SiO2 Si

e- e- e- e- e-

Si3N4 SiO2 Si
e- e- e- e- e-

5

dCTP

dATP

dGTP

T
G

dTTP

3 1 -1

C

A

C

DNA

Label-free DNA sequencing based on intrinsic molecular charge
2010/10/28








VG



Debye

100 mM ~1 nm 1 mM ~10 nm

2010/10/28



10-base 3.40 nm Change of threshold voltage, DVTdiff (mV) 20-base 6.80 nm T. Sakata et al., Biosensors & Bioelectronics 22, 1311 (2007).

DNA
30-base 10.20 nm 40-base 13.60 nm 50-base 17.00 nm

30 25 20 15 10 5 0
21-base 1 7.14 nm 31-base 2 10.54 nm 41-base 3 13.94 nm 51-base 4 17.34 nm 61-base 5 20.74 nm

DNA


2010/10/28









50/1 301x1(1 )
JST

11
2010/10/28

Protein

2010/10/28

Open Sandwich Immunoassay
Open-Sandwich (OS) Immunoassay
Ueda, H. et al. Nat. Biotechnol. 1996, 14, 1714-1718.

Sandwich assay
antigen
VH VH VL

Secondary antibody

Heavy chain

antigen
VL

Light chain

Primary antibody
Antibody

Principle of Open Sandwich Immunoassay

Impossible to bind low antigen/ VH / VL conjugate molecular antigen by the general sandwich assay!

Substrate Enzyme-VH Ag Immobilized-VL

Product

Possible merits
Detection of small antigen simple and high throughput detection
2010/10/28

Electrical detection of low molecular weight antigen using OS-FET
Open sandwich-based field effect transistor(OS-FET)
Sakata, T. Ihara, M., Makino, I. Miyahara, Y. & Ueda, H., Analytical Chemistry, in press (2009).

Ref. electrode VL VG

Low molecular antigen VL

Low molecular antigen

VH
Si3N4 SiO2
eeeee-

VH
Si3N4 SiO2
eeeee-

n-Si

n-Si

Real-time measurement Label-free Highly sensitive detection Effective method against detection of low molecule

2010/10/28

Molecular charges of VL chain
MBP(Maltose Binding Protein, MW: 42000
KIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLEEKFPQVAATGDGPDIIFWAHDRFGG YAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKLIAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELK AKGKSALMFNLQEPYFTWPLIAADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNAD TDYSIAEAAFNKGETAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKEL AKEFLENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAFWYAV RTAVINAASGRQTVDEALKDAQTGLNDIFEAQKIEWHE

Amino acids with negative charges aspartic acid D, glutamic acid E; 54 Amino acids with positive charges lysine K, arginine R; 41
MBP VL Small antigen VH
Si3N4 SiO2
eeeee-

13 Negative charges in total net

n-Si

2010/10/28

Electrical detection of low molecular weight antigen using OS-FET
Electrical signal of OS-FET
Small antigen before open
Drai current (A)
3000 2500 2000 1500

Sakata, T., Ihara, M. et al., Analytical Chemistry, in press (2009).

Bisphenol A (1mM)
710
before open sandwich after open sandwich

sandwich after open sandwich

705
700 695
0 2 4

Bisphenol A (MW228.29) 1000
500 0 -4 -2 Gate voltage (V)

13.1 mV

690 0.03 0.04 0.05 0.06 0.07 0.08

Threshold voltage (VT) Negative charge +VT (VL with MBP) (positive direction)

VG-ID electrical characteristic

Bisphenol A was detected using the OS-FET
2010/10/28

Enhancement of detection sensitivity by addition of charged molecule

MBP(Maltose Binding Protein, MW: 42000
KIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLEEKFPQVAATGDGPDIIFWAHDRFGG YAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKLIAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELK AKGKSALMFNLQEPYFTWPLIAADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNAD TDYSIAEAAFNKGETAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKEL AKEFLENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAFWYAV RTAVINAASGRQTVDEALKDAQTGLNDIFEAQKIEWHE Amino acid with amino group at the side chain lysine K

Isothiocyanobenzyl-EDTA
C18H21N3O8S=439.44

Addition of charged molecule to MBP enhances detection sensitivity of field effect-based devices
2010/10/28

Effect of addition of charged molecule on VG-ID characteristic
ICB-EDTA

710

before open sandwich after open sandwich

Drai current, mA

705

700

30.0 mV
695
Sakata, T. et al., Analytical Chemistry, in press (2009).

690 0.03

0.04

0.05

0.06

0.07

0.08

Gate voltage, V Electrical signal was amplified by adding charged molecules
2010/10/28

Detection sensitivity of OS-FET
Electrical detection of OS-FET
Threshold voltage change , DVT (mV)
35 30 25 20 15 10 5 0 -5
without EDTA with EDTA
Sakata, T., Ihara, M. et al., Analytical Chemistry, in press (2009).

control 0M

1 fM

1 pM

1 nM

1 mM

Concentration of bisphenol A

Highly sensitive detection of small antigen using the OS-FET
2010/10/28

Bodysensing (in vitro and in vivo)

QT ()



iPS



()
2010/10/28

/


- ()



()
Surface potential (mV) 9 7 5 3 1 -10 -3 -5 TRAIL K+ TNF 5 10 Time (h) 15 K K
+

Cl



/
Ag/AgCl VG Si N 3 4 SiO2 n-Si e e e e e

()
16cell



(16) FETPDMS()100mm 100mm Micro.Eng. (2008) Sakata et al. to be submitted.

FET ()

Sakata et al.

FET ()
35

(mV)

MBP

VL

VH
Si3N4 SiO2
e-

30 25 20 15 10 5 0

without EDTA with EDTA

2009-133800 Sakata et al. Analytical Chemistry (2009).

e-

e-

e-

e-

n-Si

-5

Cont.1 fM 1 pM1 nM 1 mM

pM

Subscribe
x