Fluorogenic Polydiacetylene Nanosomes: Application to Label-Free - - PDF document

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Fluorogenic Polydiacetylene Nanosomes: Application to Label-Free - - PDF document

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Fluorogenic Polydiacetylene Nanosomes: Application to Label-Free Nanobio Sensors Dong June Ahn ahn@korea.ac.kr, +82-2-3290-3301 Department of Chemical & Biological Engineering College of Engineering, Korea University Seoul, Korea April

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Fluorogenic Polydiacetylene Nanosomes: Application to Label-Free Nanobio Sensors

Dong June Ahn ahn@korea.ac.kr, +82-2-3290-3301 Department of Chemical & Biological Engineering College of Engineering, Korea University Seoul, Korea April 17-19, 2008

  • D. J. Ahn, Korea University

Diagnoses & Drug Discovery

Bio Chips & Chemical Chips

Needs for Sensors

Safety for Processed Food

Anthrax

New & Mutated Infectious Diseases Need for Disease Monitoring

Astronomical cost spent each year worldwide !

SARS Mutated Cholera, etc.

  • D. J. Ahn, Korea University
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Outline

Polydiacetylene Supramolecules Detection Examples Nanosome-Based Chips Nanosome Strips and Fibers Summary

  • D. J. Ahn, Korea University

Current Concept for Sensing Systems and Chips

  • D. J. Ahn, Korea University

Target Substance Recognizing Materials Human Judgment A B C A A A B B B C C C A A A Interface I Target / Sensor Matrix Affinity Shape Size Interface II Sensor Matrix / Human Signal Transducer Transistors / Optics Electrochemistry Mechanics / Nanowires

  • Sensor Systems

Labels: Cy5, Cy3, nanoparticles, radio tracers

Protein Chip DNA Chip

A G T T C A T C A

Pre-treatments For Labeling Analytes

  • Diagnostic Array Chips
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Why Conjugated Systems?

  • D. J. Ahn, Korea University

target molecule

Sensitivity depends on the equilibrium constant.

S T S-T

Keq = [S-T] / [S] [T]

  • Conventional Chemosensor

target molecule

Signal amplification can be achieved.

  • π-Conjugated Polymer Sensor

π-Conjugated Polymers

  • D. J. Ahn, Korea University
  • π-Conjugated Polymers

R R R R R R

S S O O O O O n NH HN O O O O O n R R R R R R N N

Polythiophene Polypyrrole Poly(phenyleneethylene) Combination (PPE + Bipyridil) Polydiacetylene

  • Electrical and/or optical properties

Easily dispersed into aqueous media !

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Polydiacetylene Supramolecules

  • D. J. Ahn, Korea University

Ligand O

Hydrophilic head molecular recognition Hydrophobic tail diacetylene unit nanosome (vesicle) solution self-assembly

X O X O X O X O X O X O X O X O X O X O X O X O

molecular spacing : 4.8±0.2Å AFM

solid-supported films Langmuir-Blodgett Langmuir-Schaefer dispersion by sonication

size : 50 nm

  • Synthetic Diacetylene Supramolecules

lipid bilayers

hydrophobic tails hydrophilic head groups inner aqueous compartment

integral protein phospholipid cholesterol hydrophobic α-helix glycolipid

  • ligosaccharide

Cytosol

  • Natural Cell Membranes

SEM

Optical Characteristics

  • D. J. Ahn, Korea University
X X X O O O X X X O O O X O X O X O X O X O X O

hν

Colorless Blue Red

X X X O O O X X X O O O

Stimuli

Real-Time Observation

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Polydiacetylene-Based Sensor Systems

  • D. J. Ahn, Korea University

conjugated p-orbital twisted p-orbital blue red target

color change

(Increased response)

Blue phase (Non-fluorescent) Red phase (Fluorescent)

Detection Examples

  • D. J. Ahn, Korea University
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increase of virus concentration

Detection of Influenza Virus A

Charych et al., Science, 261, 585 (1993); J. Am. Chem. Soc., 117, 829 (1995).

  • D. J. Ahn, Korea University

OH NH OH O O O 5 6 5 5 6 6 O O O NH O O COOH OH OH HO AcHN HO

sialic acid

N H H N N H O R1 O R2 O R3

  • Antibody: Jelinek (2001)

peptide

NH NH O O O 5 6 5 5 6 6 NH

  • Small Molecule (Cyclodextrin):

Ahn & Kim (2003, 2004)

pore match

  • DNA: Wang & Ma (2005)

Detection Examples

  • Influenza Virus A: Charych (1993, 1995)
  • D. J. Ahn, Korea University
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“Conceptual Evolution”

Films on Substrates

Method: Langmuir-Schaefer Use: Film-type Sensor Low Signal One Binding Event per Film

OH OH OH O O O 5 6 5 5 6 6

Embedded Nanosomes

Method: Polymer Blending

“Nanosome Strips” “Nanosome Fibers”

Method: Molding & Spinning Use: Chemical & Bio Sensor Blue Blue Blue Purple Pink Red

Advanced Functional Materials, 16, 2103 (2006). Advanced Materials, 19, 521 (2007).

Surface Immobilized Nanosomes Patterned Nanosomes “Nanosome Chips”

Method: Surface Chemistry

Advanced Materials, 15, 1118 (2003).

Method: Microcontact Printing Use: Sensor One Binding Event per Patterned Nanosome Surface Method: Inkjet Spotting Use: Chemical & Biological Chip Simultaneous Screening & Detection

NH 2 NH 2 NH 2 N H 2N H 2N H 2N H 2N NH 2 H N H

200㎛

Self-Emitting Fluorescence “Label-Free System”

New Concept

integration

  • J. Am. Chem. Soc., 127, 17580 (2005).
  • D. J. Ahn, Korea University

Nanosome Solutions

Method: Sonication Use: Solution-type Sensor Medium-High Signal One Binding Event per Solution

Nanosome-Based Chips

  • D. J. Ahn, Korea University
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Nanosome Array Chips

  • D. J. Ahn, Korea University

< Ink-jet Microarrayer >

Pre-spot Nanosome Library Solutions in 96 or 384 well plates A P S g l a s s

< Microarray of Nanosomes >

Inkjet (non-contact), single tip

1 spot (4 nL), width: 200 ㎛

  • Conventional sol’n analysis: 0.1 mL
  • 25,000-fold saving
  • Tiny amount of analytes
  • Simultaneous multiple screening

Microarray-Based Fluorescence Profiles

α-CD (30mM ) γ-CD (30mM) γ-CD → Heating Linear Carbohydrate : Maltoheptaose(30mM) Maltoheptaose → Heating Polyacrylic acid (30mM)

  • D. J. Ahn, Korea University

Ahn et. al., J. Am. Chem. Soc., 127, 17580 (2005).

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initial after exposure to anti-rabbit secondary antibodies after heated after heated after heated

A Label-Free Detection of Protein-Protein Interaction

when primary antibody is absent after exposure to anti-mouse secondary antibodies

  • D. J. Ahn, Korea University

Advanced Functional Materials (2007); Macromolecular Research (2006).

A Label-Free Detection of Bacteria

  • D. J. Ahn, Korea University
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Detection of E. Coli

PCDA-Aniline + PCDA-EDEA-s-NHS PCDA + PCDA-EDEA-s-NHS when primary antibody is absent

  • D. J. Ahn, Korea University

Submitted to Angew. Chem. Int. Ed. (2007).

DNA Detection Sensitivity

100 nM 10 nM 1 nM 100 pM 10 pM 1 pM 100 fM 10 fM 1 fM Spotting of vesicle-probe DNA complex Polymerization target DNA

200 ㎛

3` - GAT GGA TGC CCA TTG - 5`-NH2 5` - CTA CCT ACG GGT AAC - 3`

  • E. coli lethal factor sequence 15 mer.
  • D. J. Ahn, Korea University

100 nM 10 nM 1 nM 100 pM 10 pM 1 pM 100 fM 10 fM 1 fM

  • 50

100 150 200 250

Intensity (a.u.) Target DNA conc.

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target DNA Vesicle & probe DNA Pre spot Solution Polymerization

200 ㎛

A G T TAG Mismatched DNA

Double strand Single strand Immobilization

]

n

[

Si O O Si Si O O Si Si O O Si Si O O Si Si O O Si Si O O Si

]

n

Si O O Si Si O O Si Si O O Si Si O O Si Si O O Si Si O O Si

Filter 3. (green)

DNA Detection Selectivity: Perfect Match vs. Single-Base Mismatch

Vesicle

  • D. J. Ahn, Korea University

NH2 – 5` - GTT ACC CGT AGG TAG - 3` 3` - CAA TGG GCA TCC ATC - 5` TAG A G T

Nanosome Strips and Fibers

  • D. J. Ahn, Korea University
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PCDA liposomes in PVA

  • D. J. Ahn, Korea University

Confocal Fluorescence Microscopic Image Thermal Response Drying

Nanosome-Embedded Strip Films

O

5 7

OH OH OH O O O

5 7 5 5 7 7

OH

α-CD γ-CD β-CD control strip solution

  • D. J. Ahn, Korea University

“Litmus-Type” Strip Sensors for Chemicals

Chemistry Letters, 35, 560 (2006).

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  • D. J. Ahn, Korea University

Image Display Using Nanosome-Embedded Films

after photopolymerization thru a photomask after heating after photopolymerization

  • f the remaining area

Advanced Functional Materials, 16, 2103 (2006).

hv

electrospinning Randomly-distributed DA

R

1

R2 R2 R1 R

2

R

1

R

1

R2 R2 R1 R2 R1 R2 R1 R2 R1 R2 R1

PDA-embedded fiber Self-assembled DA

R2 R1 R2 R1 R2 R1 R2 R1

Electrospun Polydiacetylene Fibers

Advanced Materials, 19, 521 (2007).

  • D. J. Ahn, Korea University
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Visual Detection of Food Transmutation

  • D. J. Ahn, Korea University

hairtail, croaker, skate, etc chicken beef, pork

pH=8.8 pH=11.0 pH=12.3

Liquid Analytes Gas Analytes

Visual Alarm System

(Example Case: Formaldehyde)

HCHO detection

  • D. J. Ahn, Korea University
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+ Target

  • Target

Blue Red No Fluorescence Fluorescence

100㎛

Self-Emitting Fluorescence “Label-Free Chip System” Blue Blue Blue Purple Pink Red

100㎛

Color-Changing “Label-Free Strip System”

+ Target + Target

Label-Free Chips and Strip Sensors Reusable or Continuous-Monitoring System

Summary of Sensor Research

  • D. J. Ahn, Korea University

Acknowledgments

  • Dr. D. H. Charych (Lawrence Berkeley Nat’l Lab. / Chiron Corp. / Five Prime Corp.)
  • Prof. Jong-Man Kim (Hanyang University)

Research Funds from Korean Government (KISTEP, KRF, KOSEF, MOH) and Private Sectors

Home Coming Day

My Dear Students