This proposal aims to develop a novel proximity technology to quantify the hepatitis B virus (HBV) surface antigen (HBsAg) in blood for point-of-care testing. HBsAg is an important serological marker that is highly predictive of HBV infection status, and response to anti-viral treatment. Chronic infection with HBV is a serious global health problem and among the leading causes of preventable death. There are 240 million people in the world living with chronic HBV infection, with the greatest burden of disease in Asia and Africa. About 800,000 people die each year worldwide from HBV related liver disease. Improved diagnosis of HBV infection, monitoring, and treatment of HBV infected patients is urgently needed to reduce the morbidity and mortality associated with HBV infection. Risk of liver cancer is 100x higher for those who are chronically infected. WHO estimates that most of the infected have not been screened and are not aware of their infection because they often have no symptoms, and <1% of the infected that needs treatment are receiving treatment. Left undiagnosed and untreated, chronic HBV infection carries a 15-25% chance of dying from liver cancer and liver cirrhosis. Among the major challenges in scaling up prevention, surveillance, screening, care, and treatment on a global scale are affordable and easy to use diagnostic and monitoring tests, and affordable effective treatment, particularly for the resource-limited countries that have high prevalence and burden of HBV. The quantitative POC diagnostic assay we propose will have an important impact in this global effort. Specifically, we will develop a novel technology based on Fluorescence Resonance Energy Transfer (FRET) using our newly discovered quantum dot (QD) with two emissions as FRET donor, in which one emission peak without FRET serves as a ?built-in? standard and the other serves as FRET emission to interact with acceptor dye to quantify the biomarker. The assay with a feature of the ?built-in? standard could eliminate the ?matrix effect? for accurate quantification of the biomarker. This Phase I project will demonstrate its feasibility using HBsAg as a model biomarker. We will firstly prepare the QD and acceptor dye conjugates with relevant antibodies, using methods that we have established. We will then establish the quantitative FRET (QFRET) assay and study the matrix effects. The sensitivity, specificity, dynamic range, and reproducibility of the HBsAg QFRET assay will then be tested using antigen- spiked serum sample from a healthy, non-HBV infected donor. Lastly, in this Phase I study, we will evaluate the performance of the HBsAg QFRET assay by measuring the levels of HBsAg in the serum samples of HBV patients and healthy volunteers. Our QFRET HBsAg data will be compared with that of independent HBsAg assays, such as ELISA, and commercial gold standard assay for HBsAg. Our Phase II project will extend of this QFRET assay platform to quantify other biomarkers of HBV and hepatitis C virus (HCV) infections, thus providing a comprehensive solution for the diagnosis and monitoring of these chronically infected patients. Our new assay has the advantages of simple sample preparation, and a compact and inexpensive analyzer which is suitable for use in resource-limited settings.
This project aims to develop a quantum dot-based immunoassay to quantify Hepatitis disease related blood biomarkers in Point-of-Care setings. The successful development of this assay can provide rapid and accessible diagnosis of the disease, and therefore allow timely treatment intervention and monitoring of patient response to treatment. This will potentially have a significant impact on reducing the global burden of this disease.
