Lyme disease, the most common tick-borne illness in the United States, is a significant clinical diagnostic challenge. The current approach is standard or modified two-tiered tests (STTT/MTTT), which both offer limited accuracy. A biosensing platform with nanoscale plasmonic gratings to enhance signal from antibodies was developed to resolve these issues. Human serum samples with known Lyme disease status were tested on the biosensor. These samples included patients in early and late stages of the disease, as well as healthy controls. The results were analyzed with ROC analysis to determine diagnostic thresholds. Scoring the data with these thresholds showed improved accuracy over STTT and MTTT. Further machine learning analysis on the data yielded similar results to the ROC analysis.. Based on these results, this biosensor has the potential to improve clinical outcomes for Lyme disease by enabling earlier disease diagnosis through higher sensitivity toward early stages of Lyme disease.
Detection of antibodies in the blood is an important clinical technique for diagnosing active infections and previous exposures. The grating-coupled fluorescent plasmonics (GC-FP) biosensing platform has been used to detect Lyme disease serum antibodies in patients and has been shown to be more sensitive than the current standard tests. In this study, we sought to design an affordable GC-FP detection system without sacrificing the sensitivity of data output. We further optimized our analysis strategies to achieve highly sensitive and consistent diagnostic results. This work ultimately aims to fill an unmet need for better detection of human Lyme disease.
Infection with the spirochete Borrelia burgdorferi leads to Lyme disease (LD), the most prevalent tick-borne illness in the Northern Hemisphere. If left untreated, the infection spreads throughout the body, causing multisystem disease. The current standard for LD diagnosis is a two-tiered approach (ELISA followed by Western blot), which targets the immune response to bacterial proteins. This approach, however, lacks sensitivity and specificity, leading to misdiagnosis. We developed a protein microarray assay to detect antibodies against B. burgdorferi proteins with high sensitivity using grating-coupled surface plasmon resonance, combined with fluorescence imaging (Grating Coupled-Fluorescent Plasmonics, GC-FP). Here, we use GC-FP for rapid and multiplexed detection of antibodies from B. burgdorferi in human serum. We confirmed the fluorescence enhancement capability of GC-FP analysis and optimized reagent concentrations for detection of serum antibodies present in human LD. By conducting GC-FP analysis of patient serum samples, we were able to accurately diagnose LD in patients with disseminated and early-stage infection. Our results show that GC-FP can detect IgG antibodies in highly dilute human sera (up to 1:1250X serum dilution) and we are currently establishing whether or not our GC-FP platform can detect serum antibodies with greater sensitivity and specificity compared to the standard Western blot approach. Altogether, our work provides a potential path towards replacement of the cumbersome two-tiered testing algorithm, and thus a streamlined approach to LD diagnosis.
Infection with the spirochete Borrelia burgdorferi leads to Lyme disease, the most common tick-borne disease in North America, Europe, and Asia. Currently, Lyme disease is diagnosed using a two-tiered approach of ELISA/immunofluorescence, followed by Western blot analysis. These assays measure serological immune response to the infection, namely levels of IgG or IgM antibodies that bind to B. burgdorferi antigens. However, the existing approach is non-quantitative, lacks sensitivity, and may contribute to delayed diagnosis. In this study, grating-coupled fluorescence plasmonics (GC-FP) was used for rapid, highly-multiplexed detection of antibodies that bind B. burgdorferi proteins in human and mouse blood serum. GC-FP is an optical plasmonic method that enables quantitative detection of molecular interactions and can be incorporated into microfluidic format for highly multiplexed testing. We have demonstrated that this technique allows us to use only three microliters of blood serum to quantitatively detect multiple target antibodies within 30 minutes. We have also shown that GC-FP is faster and more sensitive than the traditional two-tiered Lyme disease testing scheme, making it attractive for diagnostic purposes. This proof-of-concept study provides foundations to develop GC-FP as a highly sensitive diagnostic tool to enhance the efficiency of assessment for Lyme disease patients, which will ultimately improve treatment outcomes.
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