<p>Our goals for this phase I project are:
<br>1) Improve detection limits of the paper-based assay.
<br>2) Verify Stability and Accuracy of PAD detection.
<br>3) Design and Prototype Initial Incubator/Analyzer
<br>Our objectives supporting our goals for this project are:
<p>Objective 1: Enrichment Growth Media - Time, exclusivity/Inclusivity Analysis
<br>Task 1.Optimization of L. monocytogenes and S. enterica assays with available growth media.We will test commercial enrichment broths for L. monocytogenes and S. enterica customized to fir our assay. Metrics of evaluation are: enrichment times, exclusivity and inclusivity analysis for the pathogens of interest and pathogens likely to interfere.
<br>Task 2: Validation of assays for a given food sample.Here we will test the enrichment broth against a few specific spiked food samples to determine transferability to real world samples.</p>
<p>Objective 2: Paper analytical device development and optimization
<br>Task 1. Design and fabrication of PADs for detection of L. monocytogenes and S. enterica.Paper based tests with reagents pre-loaded on paper will be developed. Preparation method, detection limits, reproducibility, geometries and form factors will be determined.
<br>Task 2. Reagent stability testing.It is a known concern that some of the molecular compounds used for bacterial pathogen detection have stability/shelf-life issues. Here we will use accelerated shelf-life testing and probe water, oxygen and temperature factors to determine the best strategy for extending and quantifying the shelf life of the paper-based devices.
<br>Task 3: Optimize spot size to minimize reagent consumption.Chemical reagent cost is a major consideration for the ultimate product. Here maximizing detection response per unit of reagent used per test will be investigated through geometric and physical variations of the test designs.
<br> Task 4: Improve response of PADs with multi-chromogen substrate incorporation. A novel approach to improving detection limits was proposed here that will explore the overlap of multiple colorimetric reagents fixed on the paper based devices. Optical absorbance and reflectance measurements will be carried out to determine if this path can lead to lower detection limits without adding complexity to manufacturing or reading results.
<br>Task 5: Validation of methods against molecular detection systems.The improved assay will be compared against gold standard methods.</P>
<p>Objective 3: Hardware Design and PrototypeThe paper-based technology will require some support from equipment that will perform timing, heating, cell disruption, and paper incubation. A rough breadboard prototype build from off-the-shelf components along with a form and fit prototype housing cut will be produced to show stakeholders and serve as the starting point for phase II productization activties.</p></p>
<p>The general scientific methods to be employed for this work are encompassed by the techniques found in microbiology, bio-analytical chemistry, analytical chemistry, materials science, mechanical engineering and electrical engineering.These efforts will be evaluated for success based on the criteria set forth in the proposal. Collecting information from the target audiences will inform and help prioritize success parameters. Determination of overall success and the success of individual tasks will happen through regular internal meetings and data review sessions. Results will be shared with external stakeholders to ensure research and development is moving toward a valuable desired product. These activities include teleconferencing and meeting in-person with our sub-awardees on this grant.</p>