Duplex Shiny app quantification of the sepsis biomarkers C-reactive protein and interleukin-6 in a fast quantum dot labe
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Journal of Nanobiotechnology Open Access
METHODOLOGY
Duplex Shiny app quantification of the sepsis biomarkers C‑reactive protein and interleukin‑6 in a fast quantum dot labeled lateral flow assay Christoph Ruppert1,2,3, Lars Kaiser1,2,4, Lisa Johanna Jacob1,2, Stefan Laufer3, Matthias Kohl1,2* and Hans‑Peter Deigner1,2,5,6*
Abstract Fast point-of-care (POC) diagnostics represent an unmet medical need and include applications such as lateral flow assays (LFAs) for the diagnosis of sepsis and consequences of cytokine storms and for the treatment of COVID-19 and other systemic, inflammatory events not caused by infection. Because of the complex pathophysiology of sepsis, mul‑ tiple biomarkers must be analyzed to compensate for the low sensitivity and specificity of single biomarker targets. Conventional LFAs, such as gold nanoparticle dyed assays, are limited to approximately five targets—the maximum number of test lines on an assay. To increase the information obtainable from each test line, we combined green and red emitting quantum dots (QDs) as labels for C-reactive protein (CRP) and interleukin-6 (IL-6) antibodies in an optical duplex immunoassay. CdSe-QDs with sharp and tunable emission bands were used to simultaneously quantify CRP and IL-6 in a single test line, by using a single UV-light source and two suitable emission filters for readout through a widely available BioImager device. For image and data processing, a customized software tool, the MultiFlow-Shiny app was used to accelerate and simplify the readout process. The app software provides advanced tools for image processing, including assisted extraction of line intensities, advanced background correction and an easy workflow for creation and handling of experimental data in quantitative LFAs. The results generated with our MultiFlow-Shiny app were superior to those generated with the popular software ImageJ and resulted in lower detection limits. Our assay is applicable for detecting clinically relevant ranges of both target proteins and therefore may serve as a powerful tool for POC diagnosis of inflammation and infectious events. Keywords: Duplex lateral flow assay, Point-of-care diagnostics, Nanoparticles, Quantum dots, Image processing, R-package, Shiny app, Sandwich immunoassay, Multiplexing, Conjugation chemistry Introduction Sepsis, a life-threatening syndrome following a dysregulated host response to infection, frequently leads to organ dysfunction; it is a major public health concern because *Correspondence: kohl@hs‑furtwangen.de; dei@hs‑furtwangen.de 1 Medical and Life Sciences Faculty, Furtwangen University, Jakob‑Kienzle Str. 17, 78054 Villingen‑Schwenningen, Germany Full list of author information is available at the end of the article
of its high mortality rates [1]. Because unspecific pathologies pose difficulties in diagnosis, the definition of sepsis has developed over time. The most recent international consensus on the definition of sepsis and septic shock, sepsis-3, was published in 2016 and defines diagnostic guidelines incl
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