2013 ColombiaUSANano

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14 Invited speakers from the US representing

Rice, Cornell, Univ. Illinois, Northeastern, UT San

Antonio, Purdue, and CTO of GE (Energy Systems)

Panelists

Carlos Fonseca, the director of Colciencias

Vice-rectors of research of U. de los Andes, U.

Nacional, Universidad EAFIT, Universidad de

Antioquia, Universidad del Valle, Universidad

Pontificia Bolivariana

Alex Orr, US embassy

Neil Vallesteros, ONR-global

Arden Bement, Purdue’s Global Policy Research

Institute

Ann Mason, then director of Fulbright-Colombia

Outcomes2013 ColombiaUSANano Univ de los Andes 2017 USAColombia Nano

Univ Antioquia- Northeastern University partnership on

nanomedicine

Uniandes-SENA-Penn State collaboration on nanotech

education

Nacional-U. Texas collaboration on nanoparticles

Sumicol Corona- Univ of Illinois collaboration on nanoceramics

Universidad Pontificia Bolivariana Nanotechnology Engineering

Program launched

SENA workshop on nanotechnology with Purdue

The Nanotechnology Center effort was been taken up by RutaN

– national policy for nanotechnology infrastructure proposed

Proyecto Interchange

2

Advances in Atomic Force Microscopy for the Characterization of Complex Materials and Live Cells

Arvind Raman

Mechanical Engineering, Birck Nanotechnology Center, Purdue University

Top ten discoveries in materials

science in the past 50 yearsMaterials Today, vol 11, 2008, P. Ball, Nature Materials, 2008

The International Technology Roadmap for Semiconductors

Scanning probe microscopes

Giant magnetoresistive effect

Semiconductor lasers and light-emitting diodes

National Nanotechnology Initiative

Carbon fiber reinforced plastics

Materials for Li ion batteries

Carbon nanotubes

Soft lithography

Metamaterials

3

4

Bio-AFM

Andor spinning disk confocal

+Asylum MFP3DCypher Asylum

Agilent 55002 Nanotec Electronica’s

Polytec LDV MSA400

Virtual Environment for

Dynamic AFM (VEDA 2.0)

AFM Research Facilities

Atomic resolution TM-AFM

mica and Calcite in water

nanoHUB resources for AFM

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D. Kiracofe, J. Melcher, S. Hu and A. Raman ~2000 users; ~15,000 simulations • Used by all major AFM companies• Air/vacuum/water simulations• AM/FM-AFM, bimodal

Virtual environment for dynamic AFM (VEDA) is a open-source, free cyber-infrastructure enabled online simulator for AFM available on nanohub (www.nanohub.org). VEDA funded by Network for Computational Nanotechnology and Dow AFM division

I have been using VEDA …

… found it to be extremely useful.

… enabled us to make better choices

in designing new probes.

… used VEDA as a check on other

calculations.

Roger Proksch, President

Asylum Research

Colombian PI’s such as Alba Avila also deploy tools on nanoHUB

AFM Trends

Quantitative physical property mapping

Combined instruments

Sub-surface imaging

High-speed imaging

Higher-resolution imaging

6

Biophysical properties of cells are

connected to biochemical state Example-Motility requires the

rearrangement of cytoskeletal networks in

the cell

Motility is driven by extension of

lamellipodia at the front of the cell and

retraction of trailing edge driven by actin

filaments and dynamic microtubules

7S. Etienne-Manneville (2013) Annu. Rev. Cell Dev. Biol. 29: 471-99.

Methods to probe live cell mechanics

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Bao,Espinosa, et al. Experimental Mechanics, 2009

AFM is unique in its capability to map cell properties on a substrate

Bacteria

Surface tension due to turgor pressure

Bending stiffness of cell wall

Visco-elasticity

Properties are heterogeneous

Applications to new antimicrobial drugs

9

Bacterianamehere.pbworks.com Mammalian eukaryotic cells

Viscoelasticity

Adhesion

Osmotic pressure

Properties are heterogeneous

Applications to early diagnosis

of cancer, understanding the

mechanical action of medicine

and fundamental cell biology

www.nih.gov: blue- microtubules, red- intermediate

Filaments , green- actin filaments

Mapping local properties of living cells

How to measure live cell mechanical properties with the AFM?

10

Force distance curves at a pointCross et al, 2007, Iyer et al Nature Nanotechnology, 2009

Point by point force distance curves (force volume)

Radmacher et al, Micron 2012

- Low resolution (~ 32 by 32) , Low speed ~ 100’s of pixels a minuteNormal vs malignant thyroid cell maps

Our approach- multi-harmonic AFM

Use high frequency, directly excited, resonant probes

scanning over sample at high speed

Observe changes in harmonics to map local mechanical

properties (viscoelasticity) – Multiharmonic AFM

Use harmonic balance to determine local properties

>10,000 pixels per minute throughput

11Raman, Trigueros, Cartagena et al, Nature Nanotechnology, 2011

i

β

Example 1: fibroblasts

12Raman, Trigueros et al, Nature Nanotechnology, 2011

Organ: Mammary gland; breast

Disease: carcinoma

Derived from metastatic site: Pleural effusion

Cell Type: Epithelial

MDA-MB-231 cell

Example 2: Carcinoma cells (~20 min)

Syk EGFPNucleusTubulin

Collaboration with Dr. R. Geahlen and M. Krisenko, Purdue Cancer Center

to understand the mechanobiology of Syk tumor suppression

No Syk

expressed

Low Syk

induction

Krisenko, Cartagena, Raman, Geahlen, “Nanomechanical property maps of breast cancer cells by multi-

harmonic atomic force microscopy reveal Syk-dependent changes in microtubule stability mediated by

MAP1B”, Biochemistry, 2014

Dr. R. Geahlen

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Boosting speed (~1 min., 256 by 256 pixels, 10,000’s pixels a minute)

Cartagena, Raman et al., NPG Scientific Reports, 2015 14

Cartagena, Raman et al., NPG Scientific Reports, 2015

Nanomechanical kinetics of Syk

knockdown on breast cancer cells

A0

EstorageEloss

d0

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3T3 fibroblasts

CO2-independent

medium, 37°C

Cantilever (BL-TR-

400PB) with a spherical

probe (5 μm diameter);

k = 0.03 nN/nm;

Scanner displacement

rate: 2 μm/s

Z-axis

Force-displacement curve

Z-a

xis

How to extract viscoelastic parameters directly from the force curves?Efremov, Raman et al, Measuring nanoscale viscoelastic parameters of cells directly from AFM force-displacement curves, in review Scientific Reports, 2017

Quantifying viscoelastic relaxation of live cells rapidly

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Combined fluorescence and AFMAFM

head

Spinning disk

confocal

(SDC)

microscope

Measuring cell relaxation during nano-indentation

by staining of actin cytoskeleton with SIR-actin

probe in MDA-MB-231 cells

X-Y

Z-Y

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Z-Y section X-Y section

AFM Trends

Quantitative physical property mapping

Combined instruments

Sub-surface imaging

High-speed imaging

Higher-resolution imaging

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Polymer based nano-composites use nano-fillers

CNT’s, graphene, Nanoparticles of silica, clay, Ti02, iron oxide

Applications: Sensors, transparent electronic applications (touch screen

displays), flexible energy storage devices (supercapacitors, batteries,

solar cells), Antimicrobial films in food packaging, Filtration/separation

membranes

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Scalable manufacturing of nanocomposites

Supercapacitor based on aligned

MWCNT/PANI composite films Lin, H.

et al. Scientific Reports 3, 1-6 (2013).

CNT thin-film transistors on a flexible and

transparent PEN substrate. Sun, D. et al.

Nature Nanotechnology 6, 156-161 (2011).CNT based bulk-heterojunction PA layer based on SWCNT

networks , Kymakis, Amaratunga, APL 80(1), 2012

CNT based electrode layer

Rowell et al. , APL 88(23), 2006

Polyimide film with dispersed graphene

micro-gravure coated on PETR2R microgravure and slot-

die system from MIRWEC

Roll to Roll

Systems

Opto-electro

nics

Drug Delivery Systems

Smart Packaging

Energy Generat

ion/ Storage

20H. C. Anderson, "Prinsessen paa Ærten”, 1835

Dispersion and connectivity of percolating networks of CNT’s

fullerenes, graphene, are difficult to judge other than by bulk

measurements. Can AFM help?

Is it possible to see

underneath the surface?

AFM for sub-surface nanometric imaging?

Electrostatic and stress fields probe

the sub-surface

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dC/dZ

kcontact

Kelvin Probe

Force Microscopy

Contact Resonance AFM

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0D: Barium titanate / polydimethylsiloxane

(BaTiO3/PDMS)1D: Single-walled carbon nanotubes /

polyimide (SWNTs/PI)

2D: Reduce graphene oxide / polystyrene

nanocomposite (RGO/PS) Nano-composite

ComponentsProperties

0D

PDMS Polymer, Insulator,

εr = 2.3 – 2.8

BaTiO3 Ceramic, ferroelectric

εr = 1200 (20°C, 1kHz)

1D

PI Polymer, insulator

εr = 4

SWNTs Semiconductor or metallic

2D

PS Polymer, insulator

εr = 2.4 – 2.7

RGO Carbon-based, conductor

2nd harmonic KPFM

Comparative studies of 2nd harmonic

KPFM and CR-AFM

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M. Cadena, et al., accepted, App. Phys. Lett, 2016

M. Cadena, A. Raman, et al. Nanotechnology, 24 (13): 135706, 2013

A. Castaneda, A. Avila, A. Raman, ACS Nano, 2014

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Computer-aided tomography

M. Cadena, et al., accepted, App. Phys. Lett, 2016

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Region 1 Region 2

Computer-aided tomography

M. Cadena, et al., accepted, App. Phys. Lett, 2016

A. Castaneda, A. Avila, A. Raman, ACS Nano, 2014

AFM Trends

Quantitative physical property mapping

Combined instruments

Sub-surface imaging

High-speed imaging

Higher-resolution imaging

26

27

E, s, h, H, Fad

=10MPa, 2 nN …

E, s, h, H, Fad

=10kPa, 3 nN …

Thank you for your attention!

Mapping local properties of viruses

Heterogeneity of material properties of viruses is due to the

intrinsic variation in the viral shells due to assembly of different

protein subunits, and due to the subsurface DNA structure

Elasticity, electrostatics, hydration forces, adhesion

Important for

Templating nanomaterials

Design virus mimmetic nanocontainers

Biophysics of viral infection 28

Eukaryotic virusesBacteriophages

CA B

25 nm

DNA layers

DNA stretchestail

pore

DNA

D

Bacteriophage ɸ29 Mature Virion

TailCollar

Capsid

29

Heterogeneities in material properties maps are

due to variations in assembly of protein

subunits in the viral capsid, collar and tail knob

and subsurface DNA structure.

Cartagena, Hernando-Pérez et al., RSC Nanoscale 2013

dAFM sub-surface imaging methods Dynamic AFM (repulsive regime)

Near surface

Needs large indentations on the surface

For soft materials

Ultrasound transmission using heterodyning

(Briggs, Kolosov, Cuberes, Tetard, Passian,

Thundat, Campbell, Arnold etc.)

Electrostatic methods

Electrostatic force gradients are modulated by local

dielectric properties

Local dielectric properties depend on sub-surface

properties

Can electrostatic/KPFM methods be used for sub-

surface imaging of nano-composites?

30Jespersen and Nygard APL, 2007; Zhao et al Nanotechnology, 2010

Sub-surface imaging

31

How deep can we see?

What limits the resolution?

Non-destructive 3D Tomography?M. Cadena, A. Raman, et al. Nanotechnology, 24 (13): 135706, 2013

A. Castaneda, A. Avila, A. Raman, ACS Nano, 2014

M. Cadena, et al., accepted, App. Phys. Lett, 2016

Layered sample analysis

Contrast can be seen down to 400nm32

33

Computational analysis

Field spreading limits resolution

Can computations be used for 3D tomography?

Interaction forces in dynamic AFM

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Each model has a small set of parameters describing physical properties

35

Finite element results for an AFM tip near a polyimide/SWNT composite sample. In (a), the computed electrostatic

force difference (F) and stiffness difference (k*) as a function of buried CNT bundle depth (radius = 20 nm, length

= 300 nm). The simulations for two tip radii are included for comparison. In (b), the FWHM calculated from the profile

across a CNT bundle as a function of CNT depth. A typical result, plotted in the inset, graphically illustrates the

higher lateral resolution produced by the CR-AFM technique

Computer-aided sub-surface

reconstruction and depth sensitivity