Paper-based assays for urine analysis

Abstract

A transformation of the healthcare industry is necessary and imminent: hospital-centered, reactive care will soon give way to proactive, person-centered care which focuses on individuals' well-being. However, this transition will only be made possible through scientific innovation. Next-generation technologies will be the key to developing affordable and accessible care, while also lowering the costs of healthcare. A promising solution to this challenge is low-cost continuous health monitoring; this approach allows for effective screening, analysis, and diagnosis and facilitates proactive medical intervention. Urine has great promise for being a key resource for health monitoring; unlike blood, it can be collected effortlessly on a daily basis without pain or the need for special equipment. Unfortunately, the commercial rapid urine analysis tests that exist today can only go so far-this is where the promise of microfluidic devices lies. Microfluidic devices have a proven record of being effective analytical devices, capable of controlling the flow of fluid samples, containing reaction and detection zones, and displaying results, all within a compact footprint. Moving past traditional glass- and polymer-based microfluidics, paper-based microfluidic devices possess the same diagnostic ability, with the added benefits of facile manufacturing, low-cost implementation, and disposability. Hence, we review the recent progress in the application of paper-based microfluidics to urine analysis as a solution to providing continuous health monitoring for proactive care. First, we present important considerations for point-of-care diagnostic devices. We then discuss what urine is and how paper functions as the substrate for urine analysis. Next, we cover the current commercial rapid tests that exist and thereby demonstrate where paper-based microfluidic urine analysis devices may fit into the commercial market in the future. Afterward, we discuss various fabrication techniques that have been recently developed for paper-based microfluidic devices. Transitioning from fabrication to implementation, we present some of the clinically implemented urine assays and their importance in healthcare and clinical diagnosis, with a focus on paper-based microfluidic assays. We then conclude by providing an overview of select biomarker research tailored towards urine diagnostics. This review will demonstrate the applicability of paper-based assays for urine analysis and where they may fit into the commercial healthcare market.

FIG. 1.

Healthcare spending and insurance coverage in the world and the United States. (a) Total healthcare expenditure per country as a percentage of each country's respective gross domestic product. W. H. Organization, see http://gamapserver.who.int/gho/interactive_charts/health_financing/tablet/atlas.html for Health Financing: Total expenditure on health as a percentage of the gross domestic product (%): 2014, 2016. Copyright 2017 World Health Organization (WHO). (b) Percentage of individuals in the United States under the age of 65 without health insurance coverage, by age group and sex (2015). Reproduced with permission from Ward et al., see https://www.cdc.gov/nchs/data/nhis/earlyrelease/earlyrelease201605.pdf for Early Release of Selected Estimates Based on Data from the 2015 National Health Interview Survey, National Center for Health Statistics. Copyright 2016 National Center for Health Statistics.

FIG. 2.

Nephron physiology for urine formation. Schematic of urine formation, consisting of filtration, reabsorption, secretion, and excretion steps. Reproduced with permission from Boundless, see https://www.boundless.com/physiology/textbooks/boundless-anatomy-and-physiology-textbook/urinary-system-25/physiology-of-the-kidneys-240/tubular-reabsorption-1174-413/ for Tubular Reabsorption. Copyright 2016 Boundless. Licensed under CC BY-SA 4.0/Additional labels added to the original.

FIG. 3.

Experimental and theoretical examples of flow through a paper-based substrate. (a) Wet-out fluid flow for uniform and non-uniform channels. The upper image shows equal fluid transport for all geometries initially, while the lower image shows the change is transport speed relative to the channel width. Wider regions have slower fluid transport to the greater cross-sectional area and the conservation of mass. Plot (at right) displays the distance traveled by the fluid front vs. the square root of time for strips A through D. Reproduced with permission from Fu et al., Microfluid. Nanofluid. 10(1), 29–35 (2011). Copyright 2010 Springer-Verlag. (b) Experimental and theoretical modeling results for fluid flow in fully wetted strips of varying widths and geometries. Reproduced with permission from Fu et al., Microfluid. Nanofluid. 10(1), 29–35 (2011). Copyright 2010 Springer-Verlag. (c) Schematic representation of the flow domain for fluid flow through an arbitrary cross-section. Reproduced with permission from Elizalde et al., Lab Chip 15(10), 2173–2180 (2015). Copyright 2015 Royal Society of Chemistry. (d) Illustration of water absorbed and retained by the cellulose fibers in unwetted paper, dependent on humidity. Above, a plot representing the fluid front travel distance vs. time; below, a plot showing the fluid travel speed vs. time. Reproduced with permission Liu et al. Appl. Thermal Eng. 88, 280–287 (2015). Copyright 2014 Elsevier Ltd. (e) Effect of humidity on imbibition in paper channels. For all papers included, there is a linear trend. Reproduced with permission from Castro et al., Microfluid. Nanofluid. 21(2), 21 (2017). Copyright 2017 Springer Berlin Heidelberg. (f) Schematic of a paper channel with wax (hydrophobic) boundaries. Paper is assumed to consist of stacked capillaries. The contact angle within/between the capillaries is less than 90°, while the contact angle adjacent to the boundary is greater than 90°, resulting in reduced capillary driving force along the boundaries. Reproduced with permission from S. Hong and W. Kim, Microfluid. Nanofluid. 19(4), 845–853 (2015). Copyright 2015 Springer Berlin Heidelberg.

FIG. 4.

Commercial rapid test examples with lateral flow assay highlighted. (a) Clockwise from upper-left: Determine™ HIV 1/2 Ag/Ab Combo. Adapted from Yetisen et al., Lab Chip 13(12), 2210–2251 (2013). Copyright 2013 The Royal Society of Chemistry. 5 Panel Drug Test Kit. (Image from HomeHealthTesting.com. OraQuick: HCV Rapid Antibody Test. Copyright Image copyright OraSure Technologies, Inc. All rights reserved. Used with permission. ImmunoCard STAT!^® E. coli O157 Plus. (b) General comparison of the functionality of direct (sandwich) and competitive lateral flow assays. Reproduced with permission from nanoComposix, see https://nanocomposix.com/collections/bioready-nanoparticles for BioReady Nanoparticles for Lateral Flow. Copyright 2017 nanoComposix (c) Example of a lateral flow assay, displaying the sample pad where the sample is loaded, the conjugate pad containing the reactive molecules, the test and control lines on the membrane, and the absorbent pad which provides capillary force to draw the fluid sample through the device. Adapted from Yetisen et al., Lab Chip 13(12), 2210–2251 (2013). Copyright 2013 The Royal Society of Chemistry.

FIG. 5.

Methods to improve the sensitivity of lateral flow assays. (a) Fluorescence-based signal enhancement by addition of fluorescent proves. Reprinted with permission from Li et al., Anal. Chem. 82(16), 7008–7014 (2010). Copyright 2010 American Chemical Society. (b) Dual gold nanoparticle (AuNP) conjugate-based signal amplification, which causes larger aggregates of the analyte at the control and test lines to improve visibility (note, silver can be deposited on top of the AuNPs to further enlarge the analytes). Reproduced with permission from Choi et al., Biosens. Bioelectron. 25(8), 1999–2002 (2010). Copyright 2010 Elsevier B.V. (c) Thermal contrast imaging to provide a quantitative readout of the presence of an analyte at the control and test lines. Reproduced with permission from Qin et al., Angew. Chem. Int. Ed. 51(18), 4358–4361 (2012). Copyright 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim. (d) Surface plasmonic resonance (SPR) for analyte detection due to analytes on the surface causing changes in the refractive index of the material. Reproduced with permission from Couture et al., Phys. Chem. Chem. Phys. 15(27), 11190–11216 (2013). Copyright 2013 Royal Society of Chemistry.

FIG. 6.

Various methods for reading quantitative detection of analytes. (a) Visual, naked eye-based analysis using calibrated colorimetric chart. Photograph shows 10 Parameter + ASC URinanlyss Reagent Strips by Xlar. (b) Smartphone-based reading using custom application and camera. Reprinted with permission from Guan et al., Anal. Chem. 86(22), 11362–11367 (2014). Copyright 2014 American Chemical Society. (c) Computational sensing using mobile light-based plasmonic chip sensors. Reprinted with permission from Ballard et al., ACS Nano 11(2), 2266–2274 (2017). Copyright 2017 American Chemical Society. (d) Surface plasmonic resonance sensing apparatus utilizing special diffraction grating structure to couple light onto a surface plasmon and disperse the diffracted light for spectral readout using a charge-coupled device camera. Reproduced with permission from Piliarik et al., Biosens. Bioelectron. 24(12), 3430–3435 (2009). Copyright 2008 Elsevier B.V. (e) Electrochemical-based reading of analytes using electrodes and handheld device. Reproduced with permission from Tang et al., Anal. Methods 6(22), 8878–8881 (2014). Copyright 2014 Royal Society of Chemistry; background image licensed under CC0 Creative Commons.

FIG. 7.

Selected fabrication methods of two-dimensional paper-based microfluidic devices. (a) Schematic illustration of fabrication by photolithography. Photolithography method used to pattern photoresist embedded in a paper substrate, which is then modified for application to bioassays. Reproduced with permission from Martinez et al., Angew. Chem. Int. Ed. 46(8), 1318–1320 (2007). Copyright 2007 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim. (b) Schematic illustration of fabrication using a flexography unit with a printing plate containing the relief patterns that form the hydrophobic regions to be printed on the paper. Reprinted with permission from Olkkonen et al., Anal. Chem. 82(24), 10246–10250 (2010). Copyright 2010 American Chemical Society. (c) Method of laser patterning, specifically paper cutting. A laser is used to cut through the paper to form channels. Reproduced with permission from Nie et al., Analyst 138(2), 671–676 (2013). Copyright 2013 Royal Society of Chemistry. (d) Schematic illustration and cross-sectional images of the fabrication of fully enclosed paper-based microfluidic devices. First, hydrophobic barriers are printed by wax printing, then the reagent zones are filled with sample dye, and a toner layer is laser printed on both faces of the prepared device to seal and protect the device. Reprinted with permission from Schilling et al., Anal. Chem. 84(3), 1579–1585 (2012). Copyright 2012 American Chemical Society. (e) A multi-pen plotter for fabricating paper-based microfluidics. Desktop pen plotter integrated with low-cost, 3 D-printed multi-pen holder, which was custom-designed to increase the throughput of the setup. Reproduced with permission from Amin et al., Anal. Chem. 89, 6351 (2017). Copyright 2017 American Chemical Society.

FIG. 8.

Three-dimensional paper-based microfluidic devices. (a) Schematic illustration of the fabrications steps of a 3 D paper-based microfluidic device constructed from the layering of photolithography-patterned paper and cut tape. Top-view and cross-sectional images show the three-dimensional flow of samples through the device, in both the horizontal and vertical direction. Reproduced with permission from Martinez et al., Proc. Natl. Acad. Sci. 105(50), 19606–19611 (2008). Copyright 2008 The National Academy of Sciences of the USA. (b) Schematic illustration of the conceptual origami paper-folding based 3 D microfluidic device. (c) Images depicting the device patterned as a single flat piece of paper, the device in its folded state, a holder for the device, and the resulting distribution of samples once the device is unfolded. Reprinted with permission from H. Liu and R. M. Crooks, J. Am. Chem. Soc. 133(44), 17564–17566 (2011). Copyright 2011 American Chemical Society.

FIG. 9.

Classification of proteins found in the urinary proteome classified based on the (a) cellular component, (b) molecular function, and (c) biological process. Reprinted with permission from Marimuthu et al., J. Proteome Res. 10(6), 2734–2743 (2011). Copyright 2011 American Chemical Society.