NIST Polymers Division Banner NIST Polymers Division Materials Science and Engineering Laboratory National Institutes of Standards and technology
NIST Polymers Division logo Side bar NIST Polymers characterization group logo NIST Polymers Electronics group logo NIST Polymers Biomaterials group logo NIST Polymers Multiphase group logo NIST Polymers Processing group logo NIST Polymers Combi group logo Side bar
 Our Publication:
 Group:

 Year:


 
 
button  HOME
button  Structure-Property Relationships in Dental Polymers and Composites
     button  Nanocomposite Dental Materials
  button  Structure-Property Relationships of Hydrogels for Dental and Craniofacial Applications
  button  The Effect of an Organogelator on Bioactive Dental Composites
  button   High-throughput and combinatorial methods for measuring the mechanical properties of dental materials
button  Combinatorial Methods for Rapid Screening of Biomaterials
  button  High-throughput Method for Determining Young’s Modulus of Polymer Blends
  button  Inflammatory Cytokine Quantification of Cell-SCK Interactions via RT-PCR
  button  Peptide Derivatized SCK Nanoparticles
  button  Real-Time Polymerase Chain Reaction
  button  Gradient Library Screening of Cell-Material Interactions
  button  Surface Energy Gradients for Characterizing Cell-Material Interactions
  button  High-throughput Method for Characterizing Cell Response to Polymer Crystallinity
  button   Cellular Response to Bis-GMA/TEGDMA Vinyl Conversion Gradients
button  Metrologies for Tissue Scaffolds
  button  Focal Adhesions of Osteoblasts on Poly(d,l-lactide)/Poly(vinyl alcohol) Blends by Confocal Fluorescence Microscopy
  button   2D -->3D Cell / Scaffold Interactions
  button  Development of a Reference Scaffold
  button   In Vitro Cartilage Development
  button   Gene Expression Profiles of Cells in Response to Tyrosine Polycarbonate Blends
  button Broadband Coherent Anti-Stokes Raman Scattering (CARS) Microscopic Imaging
  button Collinear Optical Coherence and Confocal Fluorescence Microscopies
 

line 		line  
 

Broadband Coherent Anti-Stokes Raman Scattering (CARS) Microscopic Imaging
 

Introduction
line

CARS Microscopy is a spectroscopic imaging technique which is sufficiently non-invasive to be applied to live cells. We are developing broadband CARS microscopy, which can gather roughly 10 X more spectral information than the current state-of-the-art CARS microscopy methods.
The increased spectral information we gather will position broadband CARS microscopy to provide a unique opportunity to track details of cell behavior non-invasively, and in vitro on a submicron lengthscale. Current methods that would yield similar levels of biological information are either highly invasive to live tissues or they lack the ability to image with high spatial resolution.
This method could open many exciting opportunities, including:

  • in-situ tracking of cell function for Tissue Engineering (TE) and systems biology
  • 100% QA/QC testing of TE products
  • precision histopathology
  • possibly “virtual biopsy” capabilities for cancer detection

    • Experimental Approach

    		 Broadband Coherent Anti-Stokes Raman Scattering (CARS)
    DF: dispersionless filter.
    TF: tapered fiber.
    LP: long-pass filter.
    DBS: dichroic beamsplitter.
    Obj: microscope objective
    SP: short-pass filter
    line
    CARS is a four-wave mixing technique wherein pump, probe, and Stokes light impinge on the sample, and anti-Stokes light is emitted. The key to increased CARS spectral bandwidth is an increase in the bandwidth of the Stokes light. A narrow spectral portion of output from a titanium-sapphire laser is used as pump and probe light. The Stokes light (continuum: 500 – 1100 nm) is produced by focusing the remaining laser output into a tapered fiber. Both the pump and Stokes light are combined, introduced into a high-NA microscope objective and focused on the sample. The anti-Stokes signal is collected by a spectrograph and CCD.

    Results

    line
     
    Broadband CARS spectrum Broadband CARS image Broadband CARS spectrum of benzonitrile, demonstrating the broad bandwidth of the instrument – We get sufficient bandwidth to monitor cell behavior.
    Broadband CARS image of a polymer blend consisting of polystyrene (green), polymethyl methacrylate (red), and polyethylene terephthalate (yellow). Each component is “chemically” identified with its unique broadband CARS spectrum.
     
    CARS image of adipocytes. Contrast is provided by the CH2 asymmetric stretch band at 2800 cm-1. Full spectral detection of biological samples will be possible after further noise reduction.

    Future Directions
    line Equipment, method, and analysis improvements for:
  • Reduced background, increased signal
  • Increased spatial resolution
    Application to systems biology and cell response to TE constructs and environment
    		•

    Publications
    line

  • T. W. Kee and M. T. Cicerone, “A simple approach to one-laser, broadband CARS microscopy”, Optics Letters 29 (3) 2701 (04).
  • T. W. Kee and M. T. Cicerone, “ Broadband CARS microscopy”, Microscopy Today, In Press
  • T.W. Kee and M.T. Cicerone, "A Simple Device for Broadband Compensation of Axial Chromatic Aberration," In Preparation.

    • Contributors:
    line

    Marcus T. Cicerone
    Tak W. Kee
     
     
    		 
    		 
    line
    NIST logo
    Biomaterials Group
    Polymers Division
    Materials Science and Engineering Laboratory
     
    		NIST Polymers Division Logo