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University of Ljubljana Faculty of Mathematics and Physics. Microrheology with optical tweezers. Biljana Stojković Mentor: Prof. Dr Igor Poberaj. Ljubljana, December 4th, 2012. Outline. Introduction Microrheology Optical tweezers. Passive Microrheology Active Microrheology

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slide1

University of Ljubljana

Faculty of Mathematics and Physics

Microrheology with optical tweezers

Biljana Stojković

Mentor: Prof. Dr Igor Poberaj

Ljubljana, December 4th, 2012

outline
Outline
  • Introduction
    • Microrheology
    • Optical tweezers
  • Passive Microrheology
  • Active Microrheology
  • Rheology of bacterial network
  • Future work
slide3

Microrheology

  • Rheology

Rheology is the study of the deformation and flow of a material in response to applied force.

solid

DNA

V

I

S

K

O

E

L

A

S

T

I

C

materials

properties

polymers

gels

foams

bacteria

fluid

slide4

Applying oscillatory shear strain:

Resultant shear stress:

slide5

Microrheology is

“rheology on the micrometer length scale”

  • Microscopic probe particles
  • Locally measure viscoelastic parameters
  • Study of heterogeneous environments
  • Requires less than 10 microliters of sample
  • Biological samples – limited amount of material
  • Important for fundamental reaserch and in industrial applycations
  • Current techniques can be divided into two main categories:
  • active methods that involve probe manipulation
  • passive methods that rely on thermal fluctuations of the probe
slide7

Optical tweezers technique

  • Tightly focused laser beam
  • Dielectric particles with higher refraction index that of surrounding medium
  • Wavelength of the laser  size of the object being trapped
  • Maximum force strenght is in the range of 0.1-100 pN
  • Powerful laser beam (power on sample 10 − 100 mW)
  • Microscope objective with high numerical aperture

()

slide8

How we could describe the trapping of dielectric bead?

  • R<<λ, point dipol

λ R

  • R>>λ, ray optics
slide10

Force calibration

  • Bead is held in stationary trap
  • Equation of motion:
  • Power Spectral Density (PSD):
slide11

Force calibration

  • Boltzman statistic
  • In the equilibrium, the probability density of the 1D particle position:
  • Trap potential can be obtained from normalization histogram of trapped particle postition as:
  • Fit parabola with:
slide12

Passive microrheology

  • Brownian motion
  • Two ways for determination shear modulus:

1.

Linear response theory:

2.

active microrheology
Active microrheology
  • One-particle active

Oscillations of trap:

The response of the bead is:

The equation of motion:

The viscoelastic moduli are calculated as:

active microrheology1
Active microrheology
  • Two-particle active
  • The displacements od the probe particle:
  • The same displacements can be also expressed directly as:
active microrheology2
Active microrheology

Mutual response functions:

Single particle response functions:

Complex viscoelastic modulus:

slide16

Rheology of bacteria network

Bacteria – single cell organisms

  • Different modes:
  • Free floating mode
  • Formation of biofilms
biofilms
Biofilms

Free-floating organisms attach to a surface

Colonies of bacteria embedded in an extracellular matrix (EPS)

  • EPS consist of:
  • Polymers and proteins
  • accompanied with nucleic acids and lipids
  • EPS:
  • Protect microorganisms from hostile enviroment
  • Support cells with nutrients
  • Allow comunication between cells
biofilm development
Biofilm development

Stationary phase

Death phase

Log phase

Lag phase

complexity of biofilm arises
Complexity of biofilm arises:
  • Spatial heterogeneities in extracellular chemical concentration;
  • Regulation of water content of the biofilm by controling the composition of EPS matrix;
  • Spatial heterogeneities on gene expression creates heterogeneities in polymer and surfactant production

The production and assembly of cells, polymer, cross-links and surfactants result in a structure that is heterogeneous and dynamic.

why is this study important
Why is this study important
  • Biofilm mechanics is important for survival in some enviroments
  • Well-known viscoelasticity of bioflims can provide insight into the mechanics of biofilms
  • Quantitative measure of the “strength” of a biofilm could be useful for:
    • Development of drugs for inhibition of biofilm growth
    • In identifying drug targets
    • Characterizing the effect of specific molecularchanges of biofilms.
future work
Future work

We will use optical tweezers to study viscoelastic properties of different biological samples;

  • We want to understand fundamentally how the viscoelasticity changes on different lenght scales on different frequencies;
  • Themethods willbe firsttested on water;
  • The final testground will be viscoelastic characterization of bacterial biofilms at different stages of biofilm evolution.
references
References
  • Annu. Rev. Biophys. Biomol. Struct. 1994. 23.’247-85
  • Annu. Rev. Condens. Matter Phys. 2010.1:301-322.
  • Natan Osterman,Study of viscoelastic properties, interparticlepotentials and selfordering in soft matter with magneto-optical tweezers, Doctoral thesis, University Ljubljana, 2009.
  • Natan Osterman, TweezPal – Optical tweezers analysis and calibration software, Computer Physics Communications 181 (2010) 1911–1916
  • Oscar Björnham, A study of bacterial adhesion on a single – cell level by means of force measuring optical tweezers and simulations, Department of Applied Physics and Electronics, Umeå University, Sweden 2009
  • Mark C. Williams, Optical Tweezers: Measuring PiconewtonForces, Northeastern University
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