TECHNICAL PRESENTATIONS: SESSION II

3D imaging of HIV transmission and entry 

Sriram Subramaniam, Laboratory of Cell Biology, NCI

This presentation will discuss new strategies to map viral landscapes at molecular resolution 

using novel 3D electron microscopic technologies. 

Site-Specific Chemical Mapping of Individual Cells in Two- and Three Dimensions with 

Imaging Mass Spectrometry

Christopher Szakal, Surface and Microanalysis Science Division, Analytical Microscopy Group, 

NIST

Together with collaborators from the Laboratory for Cell Biology at NIH's National Cancer 

Institute, we have begun to explore the challenging prospect of chemically mapping molecules in 

single cancer cells in two and three dimensions. By utilizing the surface sensitivity and molecular 

imaging capabilities in time-of-flight secondary ion mass spectrometry (ToF-SIMS), along with 

newly developed sample preparation protocols, we have attained state-of-the-art 400-nm 

resolution chemical maps of immortal cell lines known as HeLa cells, including the distributions 

of lipid and salt signatures. This technology can provide the foundation for exploring the sitespecific chemical changes responsible for disease progression and allow for the development of 

fast and robust imaging mass spectrometry technologies to be used in clinical settings.

Tools for Quantitative Imaging of Cells on Extracellular Matrix Mimics

John Elliott, Cell Systems Science Group, Biochemical Science Division, NIST

Quantitative fluorescence imaging and image analysis are powerful tools to measure the 

observable phenotypic characteristics of cells under experimental conditions. We have developed 

a robust two-color cell staining procedure that greatly facilitates image analysis procedures for 

determining cell morphology (structure, form and arrangement). We have also focused on the 

development of highly reproducible cell adhesion substrates coated with a fibrillar collagen type I 

extracellular matrix. The substrate preparation is compatible with many types of conventional cell 

culture plasticware and can be used in high-throughput imaging instrumentation. Our studies 

indicate that these materials mimic many properties of fibrillar collagen gels and provide 

excellent optical properties for cell imaging on an extracellular matrix substrate. These tools can 

be important components for observing phenotypic changes in cell behavior.

Live Cell Microscopy to Follow the Temporal Regulation of Gene Expression 

Michael Halter, Cell Systems Science Group, Biochemical Science Division, NIST

Quantitative measurements of dynamic processes in single cells are challenging and require the 

identification, segmentation, and tracking of live cells. Collecting and storing live-cell image data 

has been greatly facilitated by automated microscopy, but determining quantitative metrics of cell 

behavior using image-analysis algorithms remains challenging. We illustrate the application of 

live-cell microscopy and automated image analysis tools developed at NIST to measure the 

dynamics of gene expression in single cells by monitoring levels of green fluorescence protein. 

Processing for Cellular Metrology

Alden Dima, Computational Biology Project, Information Technology Laboratory, NIST

High-throughput technologies for measuring the characteristics of cells are generating large 
amounts of complex data that are difficult to process and convert into knowledge. Under the 

auspices of the NIST Computational Biology Project, experimentalists and computational 

scientists are working together to address mutually defined image-based challenge problems 

(well-defined challenges embodying essential difficulties in a research area whose solutions have 

broad impact). One challenge has been to evaluate segmentation techniques (methods to locate 

objects and boundaries in images) and associated parameters to reliably determine the cell 

morphology (structure, form and arrangement) for the purpose of comparing cell lines as part of a 

new standard procedure under development. Another challenge involves the segmentation and 

tracking of live cells in an image sequence to quantify the total fluorescence intensity of 

individual cells over time, and thereby gain a better understanding of protein expression over the 

cell cycle.

Phantom Development to Support Quantitative MRI

Robert Usselman, Biomagnetics Program, Electromagnetics Division, NIST 

NIST has recently initiated programs to support quantitative biomagnetic imaging. As part of the 

International Society for Magnetic Resonance in Medicine (ISMRM) Committee on Standards for 

Quantitative Magnetic Resonance, NIST is assisting in the design and fabrication of a new 

phantom, an object used to calibrate imaging systems. The system phantom is designed to 

measure geometric distortion, contrast properties, resolution, signal-to-noise ratio, and a variety 

of other parameters. This will be the first MRI phantom that has NIST traceability and will be 

calibrated for a range of temperatures and fields. The phantom will initially be used for quality 

control during image-based clinical trials, though widespread clinical implementation is 

envisioned. NIST is also working with the Quantitative Imaging Biomarkers Alliance (QIBA) to 

develop a dynamic, contrast-enhanced MRI phantom and will be developing susceptibility 

phantoms and flow/diffusion phantoms.

Color Contrast Agents for MRI Utilizing Magnetic Microstructures

Gary Zabow, Laboratory of Functional & Molecular Imaging (joint NINDS and NIST), Stephen 

Dodd, (NINDS), Alan Koretsky, (NINDS), John Moreland, (NIST)

A joint venture between NIST and NINDS/NIH has resulted in the development of 

microfabricated structures that can be used as MRI contrast agents with enhanced functionality or 

as micro-RFID (radio-frequency identification) tags. The microstructures can be engineered to 

appear as different effective colors when resolved using MRI as opposed to strictly grey-scale 

contrast of existing MRI agents. In this way they can be thought as radio-frequency analogs to 

quantum dots. A set of agents could be produced that would enable in vivo labeling and tracking 

of multiple different types of cells simultaneously. The agents can also act as radio-frequency 

probes of various physiological conditions. Potential applications for these structures include 

MRI, cardiovascular diseases imaging, drug development, drug candidate distribution tracking, 

diagnostics, and microfluidics.

Multicolored Fluorescent Cell Lines for High-Throughput Drug Discovery

Enrique Ubani Zudaire, Angiogenesis Core Facility, NCI

We have developed a series of immortalized cell lines that were selected to represent the different 

cell types found in angiogenesis in vivo, that constitutively express different fluorescent proteins. 

Based on these cell lines, the inventors have developed several in vitro angiogenesis assays and a 

software application that can be used to investigate the relationships between different cells 

involved in angiogenesis, to develop new combinatorial approaches to boost the efficiency of 

existing therapeutics, and to facilitate the discovery of new potential single or combination drugs. 

This technology could potentially be used to develop a high-throughput screening assay for 
angiogenesis or anti-angiogenesis drugs, or to screen compounds for cytotoxicity. The inventors 

have already demonstrated proof of concept for this technology by developing a high-throughput 

screen for potential angiogenic drugs, and they have also recently developed a cytotoxicity assay.