Theoretical analysis of immunochromatographic assay and consideration of its operating parameters for efficient designing of high-sensitivity cardiac troponin I (hs-cTnI) detection

Figures

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  Figure 1. (a) When the 99th percentile is the upper limit of the reference interval, the healthy persons in the yellow part of the distribution will get a false-positive result, and the sick persons in the red part of the distribution will get a false-negative result. (b) Illustrating that hs-cTn assays (limit set at 20 ng/L) as shown by the yellow line, detect troponin roughly 4–5 h earlier than the conventional assays (limit set at 50 ng/L) as shown by the blue line.

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  Figure 2. Distribution of troponin results for a normal population for two limits of detection (LODs) indicated by the green dashed lines. (a) LOD = 10 ng/L. (b) LOD = 5 ng/L. The 99th percentile cut points for both the assays are shown by the solid red lines.

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  Figure 3. Schematic. (i) The wall of the reactor tube (or capillary tube) is coated with labeled capture antibodies. The analyte solution is pumped into the tube. (ii) Upon entry, there is a two-way transport that takes effect: dominant advection in the streamwise (indicated by yellow horizontal arrow) and diffusion in the cross-stream (indicated by vertical purple arrow) directions. (iii) The analyte molecules displace the fluorescent labels from the sites of the antibodies. (iv) Due to consumption of the analyte at the wall and reduced transport from the middle of the tube, there is a point of exhaustion along the tube length beyond which the concentration of analyte is too low, fixed at 1 aM (10–18 M or 23.8 fg/L cTnI) in this study.

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  Figure 4. Axisymmetric concentration profile along the length of the tube at various distances (z) from the inlet. The property values are listed on the graph. These are the representative values provided in Table 2.

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  Figure 5. Influence of various control parameters on the required tube length for an immunoassay reactor. The property values are indicated on the respective graphs. Only one parameter is varied at a time for each of the four graphs. In the graphs, ‘AB’ refers to Antibody, ‘Q’ denotes the volumetric flow rate, ‘Dia’ specifies the reactor diameter and ‘\documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$${C}_{0}$$\end{document}C0’ specifies the inlet concentration. Concentration along the tube wall for different (a) inlet concentrations of cTnI, ‘\documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$${C}_{0}$$\end{document}C0’, (b) capture antibodies placed at the wall of the tube, ‘AB’ (c) volumetric flow rates, ‘\documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb}…

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  Figure 6. Concentration versus sample volume plot for typical analyte detection/analysis technologies. In the legend, m is the target number of copies (or molecules) that is to be detected per sample volume whereas, \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\lambda$$\end{document}λ is the ratio m/n or number of copies per volume/number of partitions, where n stands for the number of partitions. This graph is drawn for n = 1. The orange arrow on the concentration axis indicates the concentration level necessary for a LOD = 10 ng/L (0.42 pM) of cTnI. The horizontal region between the two blue dashed lines indicates the typical concentration range (1 nL to 1 fL) that is of interest in fluorescent immunochromatographic assays.

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  Figure 7. Multiplexed immunoassay. Here ‘n’ denotes the number of parallel channels into which the flow from a common inlet is divided. (a) Diameter of each reactor tube is \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$1 \mathrm{\mu m}$$\end{document}1μm, (b) Diameter of each reactor tube is \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$1000 \mathrm{\mu m}$$\end{document}1000μm or \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$1 \mathrm{mm}$$\end{document}1mm.

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  Figure 8. Solution of the Simple Plex™ assay on Ella Instrument. The geometric and material properties are indicated in the box. ‘\documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\mathrm{n}=4$$\end{document}n=4’ implies that the common inlet flow at \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$0.25\mathrm{ mL}/\mathrm{min}$$\end{document}0.25mL/min is divided into 4 downstream channels.