CARS Microscopy of Colloidal Gels Evangelos Gatzogiannis CARS (Coherent Anti-Stokes Raman) CARS Microscopy M.
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CARS Microscopy of Colloidal Gels Evangelos Gatzogiannis CARS (Coherent Anti-Stokes Raman) CARS Microscopy M. D. Duncan, J. Reintjes, and T. J. Manuccia Optics Letters, Vol. 7, Issue 8, pp. 350(Deuterated Onion Cells) Revived by Zubmusch, Xie in 1999. CARS Advantages •Chemical selectivity without labeling. •High sensitivity. •Signal photon at higher frequency - no spectral overlap with one-photon fluorescence background. •Small excitation volume for microscopy. •Unlike fluorescence, CARS is a coherent process and signal is proportion to ~N2 where N is the number density of scatterers. My Previous Work With CARS Elimination of Non Resonant FWM Background CARS Sample Lens pump Stokes Lens ~610nm ~575 nm probe ~610nm ~650nm ~1000cm-1 Strong vibrational resonance at ~1000 cm-1. STAND-OFF CARS SPECTROSCOPY Detector Phaseonium (Goal) Coherent Radiating Dipoles CARS Signal Epump, Eprobe x y EStokes SPORE (~1µm) DPA Molecule, i z Experimental Setup THG OPA 1kHz/10Hz Regen Stokes Tsunami UV Shaper UV CARS CARS Microscope Millenia Evolution Stokes OPA Pump/Probe OPA Quanta Ray Cost: $700,000+ Experimental Setup for RF Locking Essential for CARS, Many Uses in Metrology, Frequency Standards SHG Fs/Ps Laser 2 Delay SFG Fs/Ps Laser 1 BBO SHG 100 MHz 14 GHz 14 GHz 50 ps Phase shifter 14 GHz Loop gain 76 MHz Loop gain Phase shifter Fast Sampling Oscillosc ope SFG CrossCorrelation Laser 1 repetition rate control Stokes Laser (Master) To CARS microscope Pump/Probe Laser (Slave) 14 GHz 76 MHz Feedback Loop At the CARS Microscope, Forward vs. Epi Detection Epi-CARS Good for sub-wavelength structures, Less background Forward CARS APD/PMT Synchrolock-Based Setup ω as SFG/Cross-Correlation BBO WP/PC Pump/Probe Laser (Slave) 80 MHz 14 GHz 76Mhz Loop Gain 14 GHz Loop gain Filter WP/PC Stokes Laser (Master) Phase Shifter DBM DBM 3-D scanner 14 GHz Phase Shifter p,s Dichroic mirror as APD/PMT Simplified Setups (Improved Performance) Several CARS Images φliquid≈0.48 φxtal≈0.54 Maximum packing φRCP≈0.63 Maximum packing φHCP=0.74 Direct Imaging of Attractive and Repulsive Colloidal Glasses Repulsive Glass: Less Motion, Coop. Attractive Glass: Significant Motion J. Chem. Phys. 125, 074716 2006 Cluster Formations Is a Precursor To Colloidal Gelation – NOT Well Understood Current Experimental System This is an SEM picture of the ASM204 ~1micron colloids I am working with. Colloidal Gel Basics A gel will not form at low volume fraction unless it is buoyancy matched. For U/kBT << 1, hard sphere like behavior, monodisperse particles jam at Φ=0.63. For U/kBT >> 1, irreversible aggregation, fractal clusters are formed. Can bear stresses, have interesting mechanical properties. Physics of formation, aging, and other question remain unresolved. Most groups use fluorescence (downsides include): rapid bleaching, photo physics, alters system (in some cases, cell fixing) can’t do in vivo studies CARS: Can image for longer times (hours) depending on laser stability (without long delays frame-to-frame), Chemical specificity Resonant coherent process (better signal/background) In vivo studies, intrinsic 3d sectioning with improved spatial resolution in some cases. Noninvasive. Label-Free High Speed Imaging. No perturbation of system. Colloid-Polymer Mixtures Provide Rich Phase Behavior Blue: Gel Red: Fluid of Clusters Green: Fluids Topology and Structure Fromd 3D Images Shortest path between two particles (red stripes) along the gel, yellow, red, second shortest path. g (r ) ~ r d f 3 Radial distribution function provides direct measure of fractal dimension. Consistent with Diffusion Limited Cluster Aggregation Length of chains related to fractal dimension. Dinsmore, PRL 96 185502 (2006) Low Interaction Energies, U ≤ 2.6kBT No Structures U≥2.9kBT space filling networks with Changing Morphology Static over 30 min observation time, no signs of aging. 3D Colloidal Gel Three Days Later, After Stirring Zooming In: Colloids Still Quite Small Trajectories of Colloids In 20% Gel Van Hove Function G (r , t ) 1 N N N [r r (t ) r (0) j i 1 i j 1 G(r,t)dr is the number of particles j in a region dr around a point r at time t, given a particle i at the origin at time t=0. It separates into two terms: N Self part: 1 Gs N [r r (t ) r (0)] N Distinct part: 1 Gd N Gs(r,0) = δ(r), Gd(r,0) = ρg(r) i i 1 i N [r r (t ) r (0)] i 1 j i j i Van Hove II Signature of particles moving into positions occupied by other particles. Measures Heterogeneity and Indicative of Cage Escape