Abstract
We demonstrate a fluorescence lateral flow system that has excellent sensitivity and wide dynamic range. The illumination system utilizes an LED, plastic lenses and plastic and colored glass filters for the excitation and emission light. Images are collected on an iPhone 4. Several fluorescent dyes with long Stokes shifts were evaluated for their signal and nonspecific binding in lateral flow. A wide range of values for the ratio of signal to nonspecific binding was found, from 50 for R-phycoerythrin (R-PE) to 0.15 for Brilliant Violet 605. The long Stokes shift of R-PE allowed the use of inexpensive plastic filters rather than costly interference filters to block the LED light. Fluorescence detection with R-PE and absorbance detection with colloidal gold were directly compared in lateral flow using biotinylated bovine serum albumen (BSA) as the analyte. Fluorescence provided linear data over a range of 0.4–4,000 ng/mL with a 1,000-fold signal change while colloidal gold provided non-linear data over a range of 16–4,000 ng/mL with a 10-fold signal change. A comparison using human chorionic gonadotropin (hCG) as the analyte showed a similar advantage in the fluorescent system. We believe our inexpensive yet high-performance platform will be useful for providing quantitative and sensitive detection in a point-of-care setting.
Highlights
Lateral flow technology [1] is used for the detection of proteins, viral antigens and small molecules and enables rapid point-of-care diagnostics of infectious diseases as well as cardiac markers and cancer biomarkers.The format utilizes a sandwich immunoassay: two antibodies are bound to an analyte in a sandwich fashion
We initially began our survey with dot blots using biotinylated bovine serum albumen (BSA) spotted on nitrocellulose and detection of bound dye-labeled streptavidin
It soon became clear that the background fluorescence was limiting the sensitivity for several of the dyes, leading us to search for a quantitative approach to characterize the nonspecific binding of each of the dyes to blocked nitrocellulose
Summary
Lateral flow technology [1] is used for the detection of proteins, viral antigens and small molecules and enables rapid point-of-care diagnostics of infectious diseases (malaria [2], dengue [3,4], and HIV [5]) as well as cardiac markers (troponin [6]) and cancer biomarkers (prostate specific antigen [7]). Several reports have described the use of fluorescence in lateral flow systems [10,11,12] but their results do not show a sufficient advantage of using fluorescence instead of gold in either sensitivity or dynamic range that would justify the extra cost and complexity. We envision a new phone application that would enable on-phone data processing and reporting, providing all computer functions on the mobile device. We use this system to explore the characteristics of various fluorescent reporters in lateral flow systems. We use one of these dyes, R-phycoerythrin, in two different model systems in a lateral flow format and compare their performance to colloidal gold
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