Grantee Research Project Results
Final Report: Sensitive, Quantitative, and Portable Anatoxin Assay Using Apatamers and Quantum Dot Nanoshell Reporting
EPA Contract Number: EPD08024Title: Sensitive, Quantitative, and Portable Anatoxin Assay Using Apatamers and Quantum Dot Nanoshell Reporting
Investigators: Jackson, George W.
Small Business: BioTex Inc.
EPA Contact: Richards, April
Phase: I
Project Period: March 1, 2008 through August 31, 2008
Project Amount: $69,961
RFA: Small Business Innovation Research (SBIR) - Phase I (2008) RFA Text | Recipients Lists
Research Category: Small Business Innovation Research (SBIR) , SBIR - Water and Wastewater
Description:
Bloom-forming cyanobacteria have been observed in water bodies, including drinking water reservoirs all over the world. In addition to concern over their natural occurrence in water sources as some of the most potent biotoxins known, cyanotoxins have low-level acute toxicity that is comparable to the toxicity levels from conventional chemical weapons and, therefore, warrant consideration as a potential threat from intentional misuse (bioterrorism).
Anatoxin-a is a particularly potent, naturally occurring toxin produced by some strains of Anabaena (particularly Anabaena flos-aquae) and at least four other genera of freshwater cyanobacteria (commonly referred to as blue-green algae), including Aphanizomenon, Microcystis, Planktothrix, and Oscillatoria.
The focus of this research project is to develop a rapid assay kit for cyanobacterial anatoxins using aptamers as the primary molecular recognition element. The developed sensor chemistries will result in a detectable signal in 20 minutes or less and will require a very inexpensive, portable solid state detector system for quantitative test interpretation (readout). BioTex will first develop aptamers for highly sensitive and specific binding to anatoxins and then optimize the composition of a fluorescence resonance energy transfer (FRET)-based sensor employing quantum dot (QD) fluorescence reporters and aptamers labeled with quenching chromophores. Finally, BioTex will demonstrate the sensitivity and specificity of the novel sensing chemistry for field and laboratory use.
Summary/Accomplishments (Outputs/Outcomes):
During the Phase I effort, we have proven the feasibility of the proposed chemistry for sensing both small molecules and one small protein, insulin, as demonstrative models of our assay. Equally important, we have demonstrated our chemistry in a portable, handheld miniature fluorometer such that the assay can be taken to the field. Having optimized the sensing chemistry and detection of model analytes, we are presently selecting aptamers to the dangerous toxin and expect to have high-affinity ligands in hand for incorporation into an anatoxin-a sensor by the submission of the Phase II proposal.
Conclusions:
A novel, aptamer-based FRET affinity sensor based on quantum dots has been successfully developed. The reagents involved do not involve protein enzymes and should therefore be exceptionally stable for field use. Optimism is high for developing a mature field assay in Phase II as well as for expansion to similar analytes of concern to the U.S. Environmental Protection Agency.
Supplemental Keywords:
small business, SBIR, EPA, drinking water, water monitoring, detection techniques, measurement techniques, algal neurotoxins, cyanobacterial anatoxins, cytotoxins, drinking water system, cyanobacteria, reservoir, toxins, secondary metabolites, cyanotoxins, human health, hepatotoxins, liver damage, neurotoxins, nerve damage, anatoxin-a, saxitoxins, dermatotoxins, skin damage, Alzheimer’s, aerosolize, biotoxins, low-level acute toxicity, chemical weapons, bioterrorism, Anabaena, Anabaena flos-aquae, blue-green algae, Aphanizomenon, Microcystis, Planktothrix, Oscillatoria, rapid assay kit, aptamers, fluorescence resonance energy transfer, FRET, sensor, quantum dot fluorescence reporters, QD, quenching chromophores, sensitivity, specificity, field use, laboratory use, sustainable industry/business, scientific discipline, RFA, technology for sustainable environment, sustainable environment, environmental engineering, environmental chemistry, nanotechnology, bioterrorism, drinking water, anatoxin, biowarfare defense, point-of-use assays, portable toxin detection, biotoxin detection, bioterror agent detection,, RFA, Scientific Discipline, Sustainable Industry/Business, Environmental Chemistry, Sustainable Environment, Technology for Sustainable Environment, Environmental Engineering, biowarfare defense, bioterrorism, drinking water, nanotechnology, anatoxinThe perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Conclusions drawn by the principal investigators have not been reviewed by the Agency.