Grantee Research Project Results

2004 Progress Report: Microarray System for Contaminated Water Analysis

EPA Grant Number: R830420C004
Subproject: this is subproject number 004 , established and managed by the Center Director under grant R830420
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
Center: Center for Environmental and Energy Research (CEER)
Center Director: Earl, David A.
Title: Microarray System for Contaminated Water Analysis
Investigators: Cardinale, Jean , DeRosa, Rebecca
Institution: Alfred University
EPA Project Officer: Aja, Hayley
Project Period: September 1, 2003 through August 31, 2005
Project Period Covered by this Report: September 1, 2003 through August 31, 2004
RFA: Targeted Research Center (2004) Recipients Lists
Research Category: Hazardous Waste/Remediation , Targeted Research

Objective:

The first objective of this investigation includes developing and optimizing a novel method for glass derivatization followed by antibody adsorption to the glass surface. This will allow us to develop a multiplexed “ELISA-on-a-chip” system for antigen analysis. The second objective is to adapt our novel ELISA system for pathogen analysis in local recreational waters. The system will have the potential to fulfill a demand for a faster and more cost effective means of testing for multiple pathogens and contaminants in environmental samples.

Progress Summary:

We have hired Ashleigh Cooper, a biomaterials and engineering science graduate student, who began work on the project in September 2003. Additionally, we received a sizable donation of antibodies from U.S. Biologicals and glass substrates from Corning, Inc. We have been allowed to use the Corning, Inc. bioanalysis labs in Big Flats, New York for protein deposition and analysis.

We were able to optimize methods for chemical and UV treatment of microscope slides for use in our ELISA system. Slides of two compositions, soda lime silicate (SLS) and Corning CMT (proprietary composition), were prepared with combinations of the following different surface treatments: (1) Water plasma treatments were used to clean slide surfaces and add a thin layer of water. (2) UV treatments were performed in a Q-Sun Xenon UV and moisture control chamber. (3) Two chemical treatments were explored, 3-aminopropyltrimethoxysilane (APS) and mercaptopropyltrimethoxysilane.

Contact angles were measured to determine the effectiveness of treatments, and FTIR spectroscopy was used to determine chemical groups on the glass surface. We optimized UV treatment to 100 hours for SLS slides and 190 hours for CMT slides.

Following glass treatments, binding capacity was measured by arraying fluorescent conjugated polyclonal immunoglobulin G (IgG) on the derivatized glass surface. Initially, accurate quantification of binding was limited by a fluorescent microscope; however, recent access to a microarray printer and scanner has allowed for nanoliter printing volumes, lower testing concentrations, and higher scanning resolution. Printing, incubation, and rinsing times have been optimized in the following testing protocol:

• Slides are printed with a Genetix QArray Mini arrayer. Since the start of the project, Alfred University has acquired a microarray scanner and will have a microarray printer in February 2005.

• Slides then are incubated overnight in a humidity chamber.
• Slides are rinsed in blocking buffer with blotto to remove excess antibody and block nonspecific areas.
• Slides are dried in a centrifuge at 3,000 rpm for 1 minute.
• Slides then are viewed with an Axon Genepix scanner and images are analyzed for binding capacity with a Genepix Pro 6 program.

SLS and CMT slides were prepared with 11 combinations of the previously stated treatments and compared to three commercially available microarray slides to determine an optimal substrate for our ELISA system. Treatment using UV exposure has shown increased binding behavior for both glass substrates. UV treatment disrupts Si-O bonds, incorporates surface hydroxyls, and effectively increases surface area to allow more functionalized surface and ultimately, binding of protein. Comparison between chemical treatments indicated APS produces higher binding capacity, potentially because of the amino terminus interacting with a carbohydrate moiety or a lysine amino acid of the antibody. Preliminary data indicates that using our developed protocol, treated slides are showing higher binding capacity than commercially available slides.

Journal Articles:

No journal articles submitted with this report: View all 3 publications for this subproject

Supplemental Keywords:

ELISA, lab-on-a-chip, water contaminants, glass surface modification, microarray technology, pathogen detection,, RFA, Scientific Discipline, Ecosystem Protection/Environmental Exposure & Risk, Water, Aquatic Ecosystem, Aquatic Ecosystems & Estuarine Research, Recreational Water, Environmental Chemistry, Environmental Monitoring, recreational water monitoring, real time monitoring, microarray system, water quality, aquatic environments, risk assessment, water quality criteria, microorganisms

Relevant Websites:

http://ceer.alfred.edu/ Exit

Progress and Final Reports:

Original Abstract

Final Report

Main Center Abstract and Reports:

R830420    Center for Environmental and Energy Research (CEER)

Subprojects under this Center: (EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
R828737C001 Environmental Impact of Fuel Cell Power Generation Systems
R828737C002 Regional Economic and Material Flows
R828737C003 Visualizing Growth and Sustainability of Water Resources
R828737C004 Vibratory Residual Stress Relief and Modifications in Metals to Conserve Resources and Prevent Pollution
R828737C005 Detecting and Quantifying the Evolution of Hazardous Air Pollutants Produced During High Temperature Manufacturing: A Focus on Batching of Nitrate Containing Glasses
R828737C006 Sulfate and Nitrate Dynamics in the Canacadea Watershed
R828737C007 Variations in Subsurface Denitrifying and Sulfate-Reducing Microbial Populations as a Result of Acid Precipitation
R828737C008 Recycling Glass-Reinforced Thermoset Polymer Composite Materials
R828737C009 Correlating Clay Mineralogy with Performance: Reducing Manufacturing Waste Through Improved Understanding
R830420C001 Accelerated Hydrogen Diffusion Through Glass Microspheres: An Enabling Technology for a Hydrogen Economy
R830420C002 Utilization of Paper Mill Waste in Ceramic Products
R830420C003 Development of Passive Humidity-Control Materials
R830420C004 Microarray System for Contaminated Water Analysis
R830420C005 Material and Environmental Sustainability in Ceramic Processing
R830420C006 Interaction of Sealing Glasses with Metallic Interconnects in Solid Oxide and Polymer Fuel Cells
R830420C007 Preparation of Ceramic Glaze Waste for Recycling using Froth Flotation
R830420C008 Elimination of Lead from Ceramic Glazes by Refractive Index Tailoring
R830420C010 Nanostructured C₆B: A Novel Boron Rich Carbon for H₂ Storage