Wax-Printed Fluidic Time Delays for Automating Multi-Step Assays in Paper-Based Microfluidic Devices (MicroPADs)

Abstract

Microfluidic paper-based analytical devices (microPADs) have emerged as a promising platform for point-of-care diagnostic devices. While the inherent wicking properties of microPADs allow for fluid flow without supporting equipment, this also presents a major challenge in achieving robust fluid control, which becomes especially important when performing complex multi-step assays. Herein, we describe an ideal method of fluid control mediated by wax-printed fluidic time delays. This method relies on a simple fabrication technique, does not utilize chemicals/reagents that could affect downstream assays, is readily scalable, and has a wide temporal range of tunable fluid control. The delays are wax printed on both the top and bottom of pre-fabricated microPAD channels, without subsequent heating, to create hemi-/fully-enclosed channels. With these wax printed delays, we were able to tune the time it took aqueous solutions to wick across a 25 mm-long channel between 3.6 min and 13.4 min. We then employed these fluid delays in the sequential delivery of four dyes to a test zone. Additionally, we demonstrated the automation of two simple enzymatic assays with this fluid control modality. This method of fluid control may allow future researchers to automate more complex assays, thereby further advancing microPADs toward real-world applications.

Chronometric Quantitation of Analytes in Paper-Based Microfluidic Devices (MicroPADs) via Enzymatic Degradation of a Metastable Biomatrix

“…Our biomatrix device utilized commercially-available reagents, and therefore might be more easily adapted to a wider range of potential analytes. Our work is also similar to the published examples of passive fluid control valves (i.e., tunable delay shunts, dissolvable/erodible bridges, sucrose delays, and wax-printed time delays) [19,48,49]. However, these valves are used for assay automation and fluid control, as opposed to chronometric quantitation.…”

“…MicroPADs are typically fabricated by patterning a piece of paper with hydrophobic wax to control the flow of fluid through the porous medium via capillary action [13][14][15][16][17][18]. This increased fluid control (as compared to LFAs) allows for more complex assays to be performed on paper (i.e., multiplexed, multi-step, and/or in-series assays) [14,19,20], and many techniques have been developed for quantifying the results of these assays [21][22][23][24][25]. By far the most common quantitation technique has been the use of enzymatically-or chemically-activated colorimetric indicators, which produce a color intensity that can be correlated with analyte concentration [21,22].…”

Fabrication, Flow Control, and Applications of Microfluidic Paper-Based Analytical Devices

“…However, the disadvantage of these methods is that the paper channel or bridge is composed of chemicals, which may limit the use of certain reagents that are incompatible with the added chemical. An attempt to overcome these limitations was performed by Strong et al by fabrication of wax-printed fluidic time delays on the top and/or bottom layers of paper by varying the degree of channel coverage permitting a range of flow rates [126]. By integrating the valving functionality into wax-printed devices, a potential POC device can be created that is cost effective, simple, and reliable with the capability to conduct complex multistep assays with relative ease.…”

Predicting Dimensions in Microfluidic Paper Based Analytical Devices

“…On the other hand, flow control in paper microfluidics is still challenging, being nowadays a hot topic in research [ 32 , 33 , 34 ]. In general, liquid moves thanks to the inherent capillarity properties of paper.…”