Micro-NMR and Nanoparticle Amplification                              for Botulinum Toxin                                              Diagnostics

Recombinant Envelope Protein Domain III as a                              Candidate Subunit Dengue                               Vaccine

A Highly Sensitive, Low-labor Pathogen Detector                              Based on Retroreflector-                              linked Immunosorbent                              Assay

Genetic Screens to Identify the Ebola Virus Receptor

High-throughput Assay Development Against                              Cryptosporidium Glycotlytic                              Enzymes

Model for Oral Ingestion of Ricin Toxin

A Highly Sensitive, Low-labor Pathogen Detector Based on Retroreflector-linked Immunosorbent Assay

Collaborating Institution: University of Houston, Houston, TX

Principal Investigator: Paul Ruchhoeft, Ph.D.

Co-Investigators:
a) Robert L. Atmar, Ph.D. – Baylor College of Medicine, Houston, TX
b) Richard C. Willson, Ph.D. – University of Houston, Houston, TX

Expected Product: Micro-retroreflector-based platform technology to diagnose infectious agents.

Description: We propose to fabricate a new and very sensitive biomolecule label system and diagnostic tool based upon the extremely bright optical signals readily obtained from retroreflectors. For this proposed project, corner-cube micro-retroreflectors will be used to establish a field-compatible diagnostic for the Norwalk virus, the most common cause of viral gastroenteritis and a Group B biodefense agent. These retroreflectors consist of a transparent cube with three orthogonal, mutually touching mirrored surfaces that return incident radiation directly back to its source, making them highly detectable. In fact, a set of cubical retroreflectors on the moon's surface is routinely detected and ranged as part of ongoing experiments on lunar orbital dynamics. Retroreflectors are also used in road lane markers and bicycle reflectors (as an array of embossed cubes), but have not previously been applied to bioanalytical methods.
In our approach, we will fabricate micro-retroreflectors from glass cubes, about 5?m on a side, and coat three sides with gold. Next, the gold surfaces will be decorated with Norwalk antibodies and the cubes will be suspended into the sample solution on a sensor chip. Select areas of the surface of the sensor chip will also be decorated with stripes of monoclonal Norwalk antibodies so that, in the presence of the virus, the cubes attach to these areas on the surface. After washing, a detection step using inexpensive and field compatible detectors will probe for the bright, distinctive retroreflectance of the cubes and determine, with the aid of reference and control signals, the presence and concentration of the Norwalk virus. As a final demonstration, we will implement and validate an assay for Norwalk virus using human clinical specimens.
This work will create a low-cost diagnostic tool platform with broad applications in low-labor, inexpensive assays of very high sensitivity. With further development, the micro-retroreflector technology may potentially revolutionize diagnostics directed against NIAID Category A, B, and C agents. We expect to be able to detect the presence of single cubes, making the device extremely sensitive to even very low concentrations of pathogens.