Presentation on theme: "A revolution in micro-manipulation"— Presentation transcript:
1A revolution in micro-manipulation Optical TweezersA revolution inmicro-manipulationJonathan LeachUniversity ofGlasgow
2Today’s talk What are optical tweezers? Dynamic movement and multiple particlesCurrent research around the World
3What are optical tweezers? Optical tweezers use light to trap, manipulateand position micron sized objects.Invented approximately 20 yearsago by A. Ashkin et al.K.C. Neuman and S. M. Block, Optical Trapping, Rev. Sci. Inst., (2004)J. E. Molloy and M. J. Padgett, Lights,Action: Optical Tweezers, Cont. Phys., (2002)
4What are optical tweezers? A tightly focused laserproduces a force great enough to trap micron sized dielectric particles.Require……1. Laser2. Lens3. Object4. Damping mediumFscattFgrad
6The equipment Optical tweezers are based on high magnification microscope lensesproduces tightly focussed beamprovides visualisation of imageSamples suspended in fluidprovides coolingprovides buoyancy
7The equipment Require tight focusing so need high numerical aperture, N.A.Magnification typically X100N.A. = n sin()n is the refractive index of themedium between the objectivelens and the sample. Using oilimmersion lenses, n ~ 1.3 so N.A >1is possible.
8Optical Trapping - a>> Conditions for Mie scattering when the particleradius a is larger than the wavelength of the light .We can use a ray optics argument andlook at the transfer of momentuma
9Optical Trapping - a<< Condition for Rayleigh scattering when the particleradius a is smaller than the wavelength of the light .Scattering force and gradientforce are separableaFgrad > Fscatt requires tight focusing
10The scales Can trap 0.1 to 10’s m 1m is….. …the same as 1/100th diameter of a hair.In water, you can move a particle atabout 20-30m per sec.Require 10mW per trap.Can rotate at 100’s of Hz.
11The Q factor of optical tweezers If absorbed by particle of refractive index n, a beam of power P produces a reaction forceF = nP/c(e.g. P = 1mW: F = 5pN)The efficiency Q, of optical tweezers is defined asQ = Factual/ (nP/c)(typically Q ≈ )
12Optical Trap Dynamics Equation of motion of particle in a potential wellNewtonian forcerestoring forcedrag forceBrownian motion
13Particle in fluid Damping provided by water Solution is of exponential decay
14Particle in ideal trap Spring constant Solution is of simple harmonic motion
15Trapped particle in fluid Solution is of dampedsimple harmonic motion
16The whole picture Add in the effect of Brownian motion Time averaged effect is 0Stochastic events introducefluctuations in the particle’sposition
17Trap dynamicsLook at the movement ofthe particle in x and y
18Power spectrum Fourier transform to get the power spectrum Lorenzian Trap strength
21Exam question? In groups of 3 or 4, create two exam questions, one long, (6 marks), one short (3 marks).5mins
22Collecting data 3 options Moving 100s nm at a few kHz!!! How can we collect this data?3 options
23Option 1 - Camera Camera placed in the image plane of the objective lens.Uses the light fromthe illumination source.Fast shutter speed to takeclean image of particle.
24Option 1 - Camera Advantages Easy to use Visual image of particle Multiple particlesDisadvantages2D measurementBandwidth limitations <100HzVery slow compared to f0Require very fast shutter soneed a sensitive camera
25Option 2 - Quadrant Photodiode A Quadrant photodiode placed in the image plane of the objective lens (exactly the same as the camera).Uses the light fromthe illumination source.
26Option 2 - Quadrant Photodiode A AdvantagesLarge bandwidth 100s kHzVery fast compared to f0DisadvantagesSingle particleLow light level, so small signal2D measurement
27Option 3 - Quadrant Photodiode B Quadrant photodiode collects the laser light transmitted through the condenser lens.Small changes in the transmitted and scattered light are measured.
28Option 3 - Quadrant Photodiode B AdvantagesLarge bandwidth 100s kHzVery fast compared to f03D measurementHigh light level as collectinglaser lightDisadvantagesComplex arrangementSingle particle
30Some background optics Collimated light is brought to a focus a distance f, from a lens of focal length f.ObjectplaneImageAn angular shift in theobject plane results in alateral shift in the imageplane.
31Some background optics If the beam is notcollimated there isa shift in the axialposition of the focus.
32Moving objects aroundBeam steeringmirrorRelay lensesfffff’f’Angular deflection at mirror gives lateral shift of trap
33Diffractive optics Placing a diffractive optical element in the DiffractiongratingPlacing a diffractive optical element in theobject plane can generate a number offocused spots.
35Holograms at work split the beam focus the beam transform the beam combinations of the above
36Whole beam path also: plane waves conserved SLM SLM imaged on beam-steering mirrormicroscope objectivebeam-steering mirrormirror imaged on microscope entrance pupil
37Holographic optical tweezers can do (just about) anything! Holographic beam generation can createmultiple beamsmodified beamsfocussed beamsor all these at the same timeREAL TIME control of the beamsHologramIncident beamDiffracted beamsCurtis et al. Opt. Commun. 207, 169 (2002)
38Dynamic multiple traps Use spatial light modulator to create multiple trapsLateral displacementAxial displacementUpdate trap positionsVideo frame rateEriksen et al. Opt. Exp. 10, 597 (2002)
43Bio-applications The size of particles that can be trapped is ~0.1m to 10’s mApproximately the same size asmany biological specimen.e.g. Blood cells, stem cells,DNA moleculesEither trapped directly, or beadsused as handles to reduce opticaldamage.Ashkin et al. Nature. 330, 768 (1987)Block et al. Nature. 338, 514 (1989)
44Measuring force/motion Image trapped bead (handle) onto quadrant detectorMeasure movement of shadownm accuracy!kHz responseAdjust trap to maintain position gives measurement of forcesub-pN accuracy!biologicalobjecttrappedbeadquadrantdetectorimaging lensMolloy et al. Biophys J. 68, S298 (1995)
45e.g. Observation of myosin binding Handles attached to actin filamentIntermittent binding to myosin suppresses thermal motion of beads due to stiff physical bond
46e.g. Stretching/twisting of DNA Attach handles to ends of DNA moleculePull, let go and observe what happens!understanding of protein foldingPerkins et al. Science. 264, 822 (1994)Wang et al. Science. 282, 902 (1998)
47Work at Glasgow 5 microns Permanent micro-structures Use SLM to create tweezers arraysTrap pseudo 2D crystals (≈100) (Curtis 2002)What happens when you turn the light off?Fix structure in gelJordan et al., J. Mod. Opt.,2004
49Transfer of angular momentum Angular momentum per photon = -hbarHalf-waveplateCircularlypolarisedlightDirection ofpropagationIf the particleIs birefringent itwill absorb angular momentum and rotate.Angular momentum per photon = hbar
59A few of the (many) active groups World-wideGrier et al. NY USAGlückstad et al. Risø DenmarkRubinstein-Dunlop et al. Queensland, AustraliaUKUs! GlasgowDholakia et al. St AndrewsMolloy et al. National Institute for Medical Research, London
60Conclusions Trap dynamics and mechanisms Bio, micro, physical applicationsPositioning,manipulation andcontrol
61Constants N.A. = numerical aperture n = refractive index = angle a = radius of particle = wavelength of lightI0 = intensitynm = refractive index trapping mediumnp= refractive index particlem = np/nm (in the Fscatt, Fgrad equation)c = speed of lightm = mass (in the equation of motion)P = powerQ = trapping efficiencya = accelerationv = velocityx = positiont = timeT = temperaturekB = Boltzmann’s constantS = power spectrum= 6a= viscosity = trap strength
62Exam question? In groups of 3 or 4, create two exam questions, one long, (6 marks), one short (3 marks).5mins