Rapid point of care Hepatitis C virus detection using a long-lasting luminescent cascade signal amplification

PROJECT SUMMARY Hepatitis C virus (HCV) infection poses a formidable global health challenge, impacting an estimated 58 million individuals worldwide, with 1.5 million new infections emerging annually. If left untreated, HCV can progress to severe conditions such as cirrhosis and hepatocellular carcinoma. Despite recent strides in cost-effective HCV treatments, swift and early detection of active cases remains a key hurdle, especially in resource-limited settings. Alarmingly, only 21% of HCV-infected individuals receive a diagnosis globally, underscoring the urgent need for improved screening strategies. Of particular concern are American Indians and Alaska Natives (AI/AN), who bear a disproportionate burden of HCV infection. They experience a rate of 2.9 cases per 100,000, compared to 0.5 cases per 100,000 in African Americans and 1.2 cases per 100,000 in non-Hispanic Whites, with higher mortality rates. Achieving the World Health Organization's (WHO) HCV elimination targets, aiming for an 80% reduction in new infections and a 65% reduction in HCV-related mortality by 2030, requires substantial improvements in screening efforts. The existing two-step HCV testing protocol, involving initial antibody testing followed by confirmatory RNA testing, proves costly and time-consuming, leading to attrition in HCV management. An alternative approach lies in cost-effective, rapid, sensitive, and specific Point-of-Care (POC) HCV antigen (Ag) testing, offering potential for streamlined screening and diagnosis in a single step. However, commercially available and FDA-approved devices for POC HCV antigen testing are currently lacking. Existing assays are laboratory-based, relatively costly, and lack the necessary sensitivity and specificity, particularly for samples with low viral loads (<1000 IU/mL). To bridge this gap, there is an imperative for the expeditious development of an affordable, rapid, sensitive, and specific POC HCV Ag diagnostic assay. Here, we propose to develop POC LUCAS (LUminescence CAscade-based Sensor), an ultrasensitive bioluminescence assay designed for HCV Ag detection using a fingerprick volume of whole blood. POC LUCAS HCV Ag aims for complete automation, delivering rapid results in under 30 minutes with high sensitivity (Limit of Detection ranging from 200 IU/mL to 1000 IU/mL). This system utilizes a minimal fingerprick volume (<100 µL) of whole blood, applied to an affordable (material cost <$2), disposable, and easily mass-producible microfluidic-based cartridge. In summary, POC LUCAS HCV Ag assay stands as a promising solution to enhance access to HCV care, especially for populations disproportionately affected, providing a comprehensive and accessible approach to diagnosis in resource-limited settings and populations experiencing HCV-related health disparities.