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Nano-biophotonics for molecular imaging

Summary:

Biophotonics is at the intersection of photonics and biology where light is used to image, detect, and manipulate biological materials. This project addresses several factors that are critical for the successful development and integration of biophotonic applications in bioscience research and medicine: diagnostic and clinical standards for validating the efficacy of biophotonics-based applications, imaging standards to ensure high fidelity, a thorough understanding of the fundamental physics underlying new biophotonics-based therapeutic techniques, and measurement and science standards for improving manufacturing quality, promoting acceptance of biophotonics technology,and advancing the understanding of the interaction between biological systems and photons.

Description:

Nano-biophotonics consists of four broad areas: molecular bioimaging; nano-biosensors; multiplexed bioassays; and nanotechnology-based medical practices for diagnosis and therapy. Success in these areas is challenged by the underlying complexity of biological systems. Major levels of complexity and associated technical barriers appear at all levels of biology including the molecular, cellular, and tissue and organism levels. To address these challenges, our research strategies are based on the following approaches.

  1. Molecular level: we are developing optical measurement tools to study biomechanics of intra- and inter-molecular interactions and interactions between organic molecules and nanomaterials in a controlled environment in support of quantitative use of nanomaterials and novel probes in biology and medicine.
  2. Cellular level: we are developing optical measurement techniques to investigate molecular interactions and processes in single cells andsingle cell-based optical techniques, standards, and validation assays in support of cell-based disease diagnostics.
  3. Tissue and organism level: we are developing integrated molecular optical imaging techniques for optical diagnostics of tissues and organisms in support of standardized optical medical imaging in clinic.

Major Accomplishments:


Click here for an example of multimodality molecular imagingAt the Molecular Level:  A multimodality molecular imaging technique integrating atomic force, polarized Raman, and fluorescence lifetime microscopies, together with 2D autocorrelation image analysis is applied to the study of a mesoscopic heterostucture of nanoscale materials. This approach enables simultaneous measurement of fluorescence emission and Raman shifts from a quantum dot (QD)—single-wall carbon nanotube (SWCNT) complex. Nanoscale physical and optoelectronic characteristics are observed including local QD concentrations, orientation-dependent polarization anisotropy of the SWCNT Raman intensities, and charge transfer from photo-excited QDs to covalently conjugated SWCNTs. Our measurement approach bridges the properties observed in bulk and single nanomaterial studies. This methodology provides fundamental understanding of the charge and energy transfer between nanoscale materials in an assembly.

Areas of related measurement science and techniques include:

  • Time-correlated confocal nano-spectroscopic microscopy of single and clustered nanoprobes
  • Integrated microscopy of photo-excited nanoparticles and related processes
  • Optical tweezers and single molecule spectroscopy and microscopy

Click here for a movie of cellsAt the Cellular Level: With current concerns of antibiotic-resistant bacteria and biodefense, it has become important to rapidly identify infectious bacteria. Traditional technologies involving isolation and amplification of the pathogenic bacteria are time-consuming. In collaboration with NIH, we have achieved a rapid and simple method that combines in vivo biotinylation of engineered host-specific bacteriophage and conjugation of the phage to streptavidin-coated quantum dots. The method provides specific detection of as few as 10 bacterial cells per milliliter in experimental samples, with a 100-fold amplification of the signal over background in 1 hour. This method can be applied to any bacteria susceptible to specific phages and would be particularly useful for detection of bacterial strains that are slow growing, e.g., Mycobacterium, or are highly infectious, e.g., Bacillus anthracis. In more recent collaboration with Naval Medical Research Center, we further refined this technique for a more sensitive and quantitative assay.

Areas of related measurement science and techniques include:

  • Nano-biosensors of nanoscale building blocks
  • Multimodal optical microscopy of molecular dynamics and cellular processes
  • Hyperspectral imaging of in vivo simulated cell culture


Example screen shot of NIST's Integrated Colony Enumerator (NICE)At the Tissue and Organism Level:  Enumeration of bacterial colonies in an agar plate is simple in concept, but automated colony counting is difficult due to variations in colony color, size, shape, contrast, and density, as well as colony overlap. Furthermore, in applications where high throughput is essential, it is critical to employ a fast and user-friendly automated technique that does not compromise counting accuracy. While commercial oproducts exist that can count bacterial colonies, they can be cost-prohibitive for small laboratories. We have developed a colony counting program, NIST’s Integrated Colony Enumerator (NICE), designed to count dark colonies from multiple regions of interests on an agar plate. Images can be cost effectively acquired by a digital camera or a desktop scanner and imported into NICE. High throughput standardized assay formats such as the multiplexed opsonophagocytic killing (MOPA4) assay can be readily counted by NICE.

Areas of related measurement science and techniques include:

  • Multiplexed high-throughput biomolecular assays
  • Fusion proteins for dynamical studies of infectious diseases
  • Image analysis software for quantitative and dynamic molecular imaging
Nanobiophotonics collage

Lead Organizational Unit:

pml

Customers/Contributors/Collaborators:

Michael Emmert-Buck, National Cancer Institute, NIH
Moon Nahm, University of Alabama Medical Center
Shanmuga Sozhamannan, Navy Medical Research Center
Fuyuki Tokumasu and Thomas Wellems, National Institute of Allergy and Infectious Disease, NIH

Contractors/Guest Researchers:

Matthew Clarke
Abigail Miller

Facilities/Tools Used:

  • Time-correlated confocal nano-spectroscopic microscopy of single and clustered nanomaterials and biomolecules
  • Integrated microscopy of photo-excited nanonparticles and related processes
  • Optical tweezers and single molecule microscopy of biomolecular interactions
  • Multimodal optical microscopy of molecular dynamics and cellular processes
  • Hyperspectral microscopy of in vivo simulated cell culture
Contact

Nano-biophontics for quantitative biophysics and molecular imaging:
Jeeseong Hwang, Project Leader
301-975-4580 Telephone
301-975-6991 Facsimile

100 Bureau Drive, M/S 8443
Gaithersburg, MD 20899-8443