Superfund Research Program
Image Analysis and Bioassays Core
Project Leader: Robert C. Burghardt
Grant Number: P42ES004917
Funding Period: 2000-2008
The Image Analysis Core continues to work closely with SBRP investigators to provide routine and advanced analytical microscopy and image analysis support services. Improvement in analytical microscopy instrumentation and services including specialized training programs is an ongoing priority. New facilities added during the past year included a Zeiss Stallion Double Detector Imaging (DDI) digital imaging workstation. This workstation, co-developed by Intelligent Imaging Innovations and Carl Zeiss includes a fully motorized Zeiss Axiovert 200 MOT with fluorescence and DIC optics and a Zeiss ApoTome slider for optical sectioning. The system has been optimized for live cell imaging experiments including 2D, 2D time-lapse, 3D, rapid 3D time-lapse (4D), fluorescence resonance energy transfer (FRET), and high speed ratio imaging. During the past year, applications were added to address experimental needs of SBRP investigators who are utilizing vital imaging capabilities of the Zeiss Stallion and multiphoton microscopy instrumentation. A new Microscopy and Imaging Workshop has been developed to provide advanced training in areas of digital imaging, vital probes of cellular function, 3D imaging and specific applications with fluorescence recovery after photobleaching (FRAP), FRET and intracellular Ca2+ assessment. During the current funding period, Projects 1-3 made extensive use of vital imaging instruments (digital imaging workstations, confocal and multiphoton microscopes) and applications for routine bioassays of cellular function in toxicant-exposed cell types including: the FRAP approach to quantify effects of PAH mixtures on intercellular communication (a tumor promoter assay) and assessment of cytochrome P450 enzyme induction (EROD assay); single-cell and bulk analysis of cytochrome P450 enzyme induction; and analysis of intracellular Ca2+ homeostasis and Ca2+ waves and oscillations. Project 1 also continued utilization of immunocytochemistry as well as direct analysis of estrogen receptor (ER), aryl hydrocarbon (AhR), Sp1-Sp4 and coactivator protein-protein interactions with FRET assays. Other analyses involved the evaluation of the efficacy of specific siRNA constructs on the expression and function of these proteins. Project 4 utilized transmission electron microscopy facilities to investigate ultrastructural changes and apoptosis in the neural tube of fetal mice exposed to arsenic.