takehiko kitamori the university of tokyo -...

Takehiko Kitamori

The University of Tokyo

Micro Unit Operations (MUO) Continuous Flow Chemical Processing

(CFCP)

+

Condensation Distillation

Column Membrane

L/L

G/L

L/S

Others

Phase contact

Extraction

Phase separation

Mixing & reaction

Reaction Extraction

Concentration

Babble sep.

Adsorp. or

reac.

Heating Cell culture

Phase cont. Phase sep.

Micro Unit Operation and Continuous Flow Chemical Processing

MUO & CFCP

Sample

Aq./Org. (shaking)

Colorimetry

Organic solution

Unit operation MUO Experimental procedure

Reagent

Aq. Org.

Aq. Org.

Aq. Org.

HCl

NaOH

Aq. Org.

Water

Extraction

Phase separation

Detection

Mixing & reaction

Phase contact

Phase contact

Phase contact

Phase contact

Phase separation

Phase separation

Phase separation

・Continuous Flow Chemical Processing (CFCP)

100mm

Design Protocol for Unit Operation Based Microfluidics Example: Wet Analysis for Trace Metal

Stabilization of Parallel Microflows

Org.

Aq.

Aq.

Aq.

Org.

Aq.

Org.

Aq.

500 µm

70 mm

30 mm

140 µm

Stabilization of multiphase microflow

Aq. Org.

① Structure of channel

② Surface Modification

③ Flow condition

Format Change between Parallel-Droplet Microfluidics

By Gucha-Gucha Chip

500 µm 500 µm 500 µm Hydrophilic surface

Hydrophobic surface

Phase separation Droplet formation

Water

Oil

Hydrophilic surface

Droplet-to-plug

Hydrophilic surface

Hydrophobic surface

Our Concept of Micro Integration of Chemical & Biological Process

Micro Unit Operation

50mm

Separation Confluence

Mixing, reaction Extraction

Continuous flow chemical processing (CFCP)

Tokeshi (Kitamori Lab.), Anal. Chem., 2002

CPU

Chemical CPU

Chemical CPU

Chemical instruments

etc., etc., almost 20 kinds

Microchips Installed Chemical Instruments

m-Extraction chip and

environmental analysis system m-ELISA chip and serum

immunoassay system

Compact

Rapid

Simple m-Gas extraction chip and clean room gas monitor

7,500 parallel CPUs

Gel particle plant

(30t/year)

Institute of Microchemical Technology

IMT Products

Micro Chemical Chip Peripheral Devices & Accessories

Detectors: TLM Systems

www.i-mt.co.jp

1 ~Å nm µm mm

10 100 1 10 100 1 100 10 1

Micro chemical chip

(Device)

System

Nanotech

Quantum effect

Object Molecule,

Nano molecule

Field

Principle

CNT, Nano pore Method

Extended-nano space

Nano space

Micro chemical chip

Microspace

Continuous fluid

Micro chemistry

Classical dynamics

Bulk space

ガラス器具

Components： Micro unit operations

Circuit： Continuous flow chemical processing (CFCP)

Micro unit operations

Macro unit operations (MUO)

Mixing Confluence Extraction Separation

100 µm

10cm

Divided by 500

100nm

100nm

Extended-nano space

50 mm

UV-Vis light

Background: Micro and Extended-Nano Fluidics

Olympic Game in Analysis

Sample volume: Smaller

Concentration: Lower

Separation: Finer

Duration: Faster

Throughput: Higher

Cost: Lower

Why extended-nano fluidics?

Single Molecule Immunoassay at Femto-Liter

on Extended–nano Fluidic Device:

A Crazy ELISA (ITP 2013 Plenary)

Strategy of Micro/E-nano Fluidics for Analysis

Our world

fluidics

Micro

fluidics

Extended-nano

fluidics

Micro

fluidics

Our world

fluidics

mm/nm

interface

nm/mm

interface

Cell process Molecule process

Detection

MENU

Fabrication

Detection

Separation 1 Chromatography

Separation 2 Immunochemical

Separation 3 L/L extraction

Sampling interface

Nanofabrication (yellow room in Kitamori Lab.)

Plasma etching Glass bonding @ RT

Electron beam lithography

Substrate

EB

Development of channel fabrication method

Top-down fabrication by electron beam (EB) lithography and plasma etching

Plasma

Resist

Channel

EB & plasma etching

Optimization of EB

exposure time (0.4 ms)

100 nm

100 nm square

Extended-nano channels EB spot

100 nm

100 nm

Extended

nanochannel

Top view

Roughness: 3 nm

100 nm

Cross-section view

Depth:100 nm

Width:106 nm

Bonding

・ Weaker bonding than thermal one ・ Good for fnctionalization

Low temperature (25-100℃), several hours Pressure 1000-5000 N

Low-Temperature Bonding Giving Chemical Function to EN Channel

Thermal fusion bonding Low-temperature bonding

Catalyst

Electrode

Biomolecules

1060℃, 6 hours

Thermal fusion Burned out

・ Strong and clean bonding ・ Difficult to functionalize

Catalyst

Electrode

Biomolecule

Chemically-

functionalized

Activation by O2 plasma

Hydrophobidized by Fluorine

Surface treatment

MENU

Fabrication

Detection




Sampling interface

Fluorescence

Detection in Extended-Nano Channels (DIC: Differential Interference Contrast TLM: Thermal Lens Microscope)

Excitation beam

EN channel

d=100nm

Probe beam

phase

Phase change

EN channel

d=100nm

Change of RI

LIF DIC-TLM

For fluorescent analyte For non-fluorescent analyte

Heat

Peak a

rea[a

.u.]

0 5 10 15 20 25

0

100

200

300

400

500

600

0 5 10 15

Concentration [pM]

Absolute number [molecules]

Peak area plot

Results: Calibration Curve

0

2

4

6

8

10

12

14

16

18

0 30 60 90 120

Time [sec]

Therm

al le

ns s

ignal [m

V]

Blank

2.7 pM

5.3 pM

11 pM

Raw data

MENU

Fabrication

Detection




Sampling interface

Atto-Litter Chromatography

910 nm

470 nm

1600 mm

loading channel

separation channels

depth : 200 nm

pressure controller

valve

fluorescence microscope

Glass microchip vial

Schematic image of nanochannel

7 cm

3 cm

nanochannels

hole

microchannel

Glass or quartz microchip

15 cm

Anal Chem (2010), Small (2012), J. Chro. A (2011)( 2012)

550 aL

Normal phase

Time [s]

強度

[a

.u.]

0

0.4

0.8

1.2

0 1 2 3 4 5

Reversed phase

0

0.4

0.8

1.2

0 4 8 12 Time [s]

Hydrophilic interaction

0 1 2 3 4 5

0

0.4

0.8

1.2

Time [s]

Various Separation Modes by Surface Modification

Time [s] 0 500 400 300 200 100

0.4

0.8

1.2

0

Column HPLC

強度

[a

.u.]

0

0.4

0.8

1.2

Time [s] 0 100 200 300 400 500

Column HPLC Extended-nano

Time [s] 500 400 200 0

0.4

0.8

1.2

0 300 100

Column HPLC Extended-nano Extended-nano

Separation performance

Column HPLC :

45,000 plates/m


Nonpolar

Stationary

Stationary

Mobile

OH OH OH OH OH OH

Polar

OH OH OH OH OH OH

Polar


Polar

OH OH OH OH OH OH

OH OH OH OH OH OH

Polar

Polar

Hydrophobic interaction

Polar

CH3 CH3 CH3 CH3 CH3 CH3

Nonpolar

CH3 CH3 CH3 CH3 CH3 CH3

Nonpolar


Extended-nano :

440,000 plates/m


Column HPLC :

43,000 plates/m


Column HPLC :

44,000 plates/m


Extended-nano :

350,000 plates/m


Extended-nano :

910,000 plates/m

H = A + B/u + C・u

A

B 200 nm

C 200 nm

200 nm

No vortic diffusion

No particle

H = A + B/u + C・u

Small space

100 mm

Plate height：H

(Term A、B、C : factors to decrease separation efficiency)

Conventional HPLC Extended-nano chromatography

Vortic diffusion

Diffusion in flow direction

Diffusion in depth direction

Flow rate:u

Theoretical Discussion about Innovative Performance

MENU

Fabrication

Detection




Sampling interface

Single molecule detection and counting T

LM

sig

na

l [m

V]

Time [sec]

Detection point ：1.5 mm downstream

Concept of Single Molecular Detection in Extended-nano ELISA

200nm

2mm

Antibody-modified region： 3 mm 1.5 mm

Injection

Antibody-modified region

Non-specific adsorption

of HRP-antibody

Spot size

1.1mm

DIC-TLM Analyte in proximity Isolated analyte

Non-specific adsorption

of HRP-antibody

Diffusion length

±400 mm

1 molecule

2 molecules

Perfect capture

Results of Single Molecule ELISA

5

4

3

2

1

02520151050

secTime [sec]

TLM

sig

na

l[m

V]

Measurement 1

Measurement 2

Measurement 3

Results

Inlet Outlet

Signal from antibody-immobilized region

5 molecules

1 molecule Non-specific

Design and fabricated device

Extended-nano ELISA chamber

Inlet Outlet

Antibody

140 mm

3.3 mm

200 nm

Antigen

~100 fL

200 nm

depth

Diffuse in

~2 sec

Reaction field volume : = 100 fL

Concentration : = 10 pM (1 molecule/100 fL)

Reaction time : = 2 sec (traveling time)

Collision frequency

37000 times

DIC-TLM

MENU

Fabrication

Detection




Sampling interface

Concept of Unit Operation in Micro/Extended-nano Fluidics

Microchemistry (10-100 mm)

Integration concept using multiphase flow

50mm

Micro unit operations (MUO)

Separation Confluence

Mixing, reaction Extraction

Continuous flow chemical processing (CFCP)

Various complicated chemical process

Environmental analysis, synthesis etc.

Tokeshi (Kitamori Lab.), Anal. Chem., 2002

Aq. Org.

10-1000 nm

Extended-nano chemistry (10-1000 nm)

1/500

Novel functional devices:

Volume: aL-fL 100kPa

g : surface tension

d : diameter d

g

PLap＝

Glass

Partial surface modification

MENU

Fabrication

Detection




Sampling interface

Advantage

Proposal of Micro/Extended-nano Interface by Lipid fusion

Extended-nano channel

20 mm

900 nm

Lipid bilayer

Cell

900 nm

Lipid bilayer

Lipid fusion

(molecular interaction)

(1) Formed ~ 102 nm hole on cell membrane

Realizing micro/extended-nano sampling interface by lipid fusion

(3) Tight junction ( no leakage) by lipid fusion (2) Keeping viability of cell due to the small size

10 mm Pressure

Evaluation of Sampling Volume

L: 50 ± 0.4 μm

20 μm

50 ± 0.4 μm

Nano channel volume: 39 fL ( 978 nm × 826 nm × 50 μm )

Evaluation of sampling volume

Observation after bonding

Why Extended-nano Fluidics?

Molecular behavior restriction

Smaller than unit diffusion distance of single molecule

Scientific uniqueness

Property change

Solution properties

Fluid characteristics

Transport phenomena

Photonic properties

High viscosity (x5) dielectric cons. (x1/4)

Surface slipping

High proton mobility (x20)

Blue-shifted optical near field

Engineered uniqueness

Molecular position control

Nano fabrication and nano surface modification

= Guiding target molecule to designed position

virus Cell

femto liter ( 1 mm)3

Smallness

nano liter (100 mm)3 atto liter (100 nm)3

pico liter ( 10 mm)3 zept liter ( 10 nm)3 Bacterium

fL

200nm

Microtiter plate

Positioning in microtiter plate & extended-nano fluidic device

Extended-nano fluidic device

I am here. Where shall I be caught?

mL

1H-1/T1

Intermolecular

Intramolecular

1H experiment

1H-1/T1 OH / D

850nm

750nm

850nm

950nm

DE

Restriction of

molecular mortion

Activation energy of

proton

Proton localization Proton transfer

NMR Measurement of Water in Extended Nano-Channels

2 µm

SEM image

Anal. Chem. (2002)

98 99 00 01 02 03 04 05 06 07 08 09 10 11 120

20

40

60

80

100

98 99 00 01 02 03 04 05 06 07 08 09 10 11 120

50

100

150

200

250

300

350

Field of nanochemistry and nanofluidics

Nanopore: ~2000 papers Nanochannel: ~350 papers

Year (-) Year (-)

Num

ber

of

paper

(-)

Hibara, Kitamori, et al

Anal. Chem. (2002)

Size-regulated fluidic channel Unregulated nanospace

Si O Si O Si O Si O

Si O Si O Si O Si O

50 nm

Wall

H+

H+

H+

H+

H+

H+ H+

H+

Fluid dynamics in extended nanospace

Extended nano physical chemistry

Stokes-Einstein Hopping

2 26

B B HS H

k T k TD

r z F

m

• NMR

• Streaming potential

• Capillary action

• Electric

• nano-PIV

• STED

• RAMAN

Measurement

• viscosity (×4) • dielectric constant （×1/4） • proton mobility (×20) • conductivity (×80) • condensation T (120ºC)

• chemical equilibrium shift

• Surface Slipping flow

• High E near field

Properties

Publications • web-site of Kitamori-Lab (#>30)

EN-channel

Priventing H2,O2 mixing

O2

EN--channel （hydrophilic）

H2

H2/O2 generation and separation by light illumination

H2 O2

e-

H+ H2O

H+ transfer

H+

Nanochannel

20X Higher H+ mobility

H+ H+ Glass

Gas-liquid separation

H2

Hydrophobic

Laplace pressure to gas Laplace pressure to water

Pt cathode Nano-structured photocatalyst (photoanode)

Hydrophobic Hydrophobic

Design of Fuel (H2/O2)) Supplier Device

Automatic H2/O2 generation and separation

Pt cathode

Nanochannels

Results

500 mm 4x

Stoichiometric H2/O2 generation

WO3/BiVO4 photocatalyst

1. Observation of H2/O2 generation

2. Analysis of generated gas by GC-MS

H2 O2

0

50

100

H2 O2 Genera

tion r

ate

[nL/m

in] Generation rate of O2 : H2≒1:2

Microscope

Experimental set up

photoanode Pt cathode

Electrolyte: 0.5 M NaClO4 Bias : 0.25 V vs Ag/AgCl ref. electrode

Water separation and H2/O2 generation by Vis light illumination

Fuel (H2/O2)) Supplier Device

500nm

WO3/BiVO4 Photocatalyst

Solar Light Driven μ-Fuel Cell Device

Introduction Microchannel

(w: 600 um, d: 6 um)

PA:

WO3/BiVO4

C: Pt

Fuel Supplier (FS)

Solar Light

e-

Fuel Cell (FC)

Device Design

H2O Circulation

Nanochannels

(φ: 200 nm)

Extended-nano channels

(φ: 200 nm)

Hydrophobic channel

(w: 400 um, d: 0.5 um)

PA-photoanode; C-cathode: A-Anode

C: Pt

A: Pt/Pd

Fuel generation & separation

H2

O2 e- H2O H+

H2

O2

FC electricity generation

H2

O2

e-

H2O

H+

Nanochanel for H2O circulation

H2O

Resistance~1200MΩ: (Electrically disconnected)

Kitamori Lab 2015

http://park.itc.u-tokyo.ac.jp/kitamori/

We always welcome visitors who use our technologies

takehiko kitamori the university of tokyo -...

Documents