Ultrasensitive Detection of Hepatitis Viruses by Immunoassay Amplification

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DESCRIPTION (provided by applicant): This proposal responds to the action plan for combating the Silent Epidemic of Viral Hepatitis recently launched by the National Institutes of Health by focusing on early diagnosis of hepatitis, which is crucial for preventing further infections and to improve therapy. Approximately 170 million people are infected with HCV, many of whom develop cirrhosis and liver cancer. A large proportion of chronically infected individuals do not exhibit symptoms, thus are unaware of their risk of developing cancer, highlighting the need for comprehensive surveillance. Diagnosis during the early stages of HCV infection is currently done using nucleic acid amplification testing (NAT)-the gold standard. However, NAT is not suitable for surveying large populations or for point of- care (POC) testing because it is expensive and requires costly equipment and trained personnel. BCR Diagnostics, Inc. (BCR) proposes a cost-effective alternative to NAT by applying its innovative biosensor technology to reduce the low detection limit (LDL) of bead-based enzyme-linked immunosorbent assays (ELISAs) for HCV core protein (cAg). BCR technology exploits phenotypically engineered fluorogenic bacterial spores (F-spores(tm)) as nanodetectors. The proposed biosensor is based on an existing biochip (termed 80K-bioChip(tm)), which is a disposable device containing 80,000 independent biosensors (termed micro-colanders(r)) that are each filled with about 200 F-spores. A feature of the 80K-bioChip is the use of an inexpensive digital camera as a quantitative readout. The project has the following specific aims: Specific AIM 1: To validate the BCR biosensor for measuring low levels of HCV core protein (cAg) in human serum. Approach: The biosensor performance will be tested using human serum spiked with different concentrations of cAg. Milestone: Data showing that the biosensor significantly improves the LDL of a conventional ELISA for cAg. Specific AIM 2: Measure sensitivity and specificity of the biosensor using HCV seroconversion panels. Approach: The biosensor will be used to measure cAg levels of individual sera in a commercial seroconversion panel from patients at various different stages of HCV infection. Milestone: Data showing sensitivity and specificity near 100%. Phase II plan: To convert the biosensor into a marketable prototype for research and clinical laboratories, and to use the prototype for a study designed to obtain 510 (k) clearance from the Food and Drug Administration (FDA). Project Description