Rapid Detection of Cat Cystatin C (cCys-C) Using Immuno-Pillar Chips.

We demonstrated a rapid immunoassay for detection of cat cystatin C (cCys-C) which is an important marker for chronic kidney disease in cats, using immuno-pillar chips. The required amount of reagent solution is 200 times smaller than that for the conventional ELISA in the 96-well microplate (0.5 µL versus 100 µL). In addition, the total assay time in the proposed method is more than 12 times shorter than in the conventional method (20 min versus 240 min). The limit of detection in the new method of 3 ng mL-1 is comparable to that of the conventional method (1 ng mL-1) and it is in the clinically relevant range.

A competitive immunoassay system for microfluidic paper-based analytical detection of small size molecules.

The development of a competitive immunoassay system for colorimetric detection on microfluidic paper-based analytical devices (µPADs) is reported. The µPADs were fabricated via photolithography to define hydrophilic flow channels and consisted of three main elements: the control and test zones, where target detection was performed, the sample introduction zone, and the competitive capture zone located between the sample introduction zone and the test zone. The chromogenic substrate 3,3',5,5'-tetramethylbenzidine (TMB) was deposited at the control and test zones. µPAD surface modification was performed at the capture zone first via chitosan activation, then the BSA-conjugated target compound was immobilized. The sample solution consisting of the target compound, the peroxidase-conjugated antibody, and the hydrogen peroxide oxidizing agent was introduced into the device and competition occurred at the capture zone, allowing only the target-bound peroxidase-conjugated antibody to travel past the capture zone and into the test zone via capillary action. The developed competitive immunoassay system was successfully demonstrated on the µPAD detection of biotin as a model compound with a detection limit of 0.10 µg mL-1. The applicability of the proposed immunoassay system for point-of-need testing was further demonstrated using aflatoxin B1, a highly toxic foodborne substance, with a detection limit of 1.31 ng mL-1. The µPAD with the competitive immunoassay format showed promising results for practical applications in point-of-need testing involving small molecular weight targets in food and water safety and quality monitoring, environmental analysis, and clinical diagnostics.

Image analysis for a microfluidic paper-based analytical device using the CIE L*a*b* color system.

The combination of a microfluidic paper-based analytical device (µPAD) and digital image analysis is widely used for quantitative analysis with µPADs because of its easy and simple operation. Herein, we have demonstrated a quantitative analysis based on multiple color changes on a µPAD. The CIE L*a*b* color system was employed to analyse the digital images obtained with the µPAD. We made pH measurements using a universal pH-indicator showing multiple color changes for various pH values of aqueous test solutions. The detectable pH range of this method was wider than the typical grayscale-based image analysis, and we succeeded in the measurements for a wide pH range of 2-9.

Novel concept of washing for microfluidic paper-based analytical devices based on capillary force of paper substrates.

A novel washing technique for microfluidic paper-based analytical devices (µPADs) that is based on the spontaneous capillary action of paper and eliminates unbound antigen and antibody in a sandwich immunoassay is reported. Liquids can flow through a porous medium (such as paper) in the absence of external pressure as a result of capillary action. Uniform results were achieved when washing a paper substrate in a PDMS holder which was integrated with a cartridge absorber acting as a porous medium. Our study demonstrated that applying this washing technique would allow µPADs to become the least expensive microfluidic device platform with high reproducibility and sensitivity. In a model µPAD assay that utilized this novel washing technique, C-reactive protein (CRP) was detected with a limit of detection (LOD) of 5 µg mL-1. Graphical Abstract A novel washing technique for microfluidic paper-based analytical devices (µPADs) that is based on the spontaneous capillary action of paper and eliminates unbound antigen and antibody in a sandwich immunoassay is reported.

3,3',5,5'-Tetramethylbenzidine Oxidation on Paper Devices for Horseradish Peroxidase-based Assays.

We report on the colorimetric oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) by hydrogen peroxide using horseradish peroxidase on photolithography-fabricated (P-PAD) and wax-printed (W-PAD) paper-based analytical devices. Fabricating PADs via photolithography exposes the hydrophilic areas to polymers (photoresists) and solvents, not only reducing the hydrophilicity, but also affecting the TMB-H2O2 assay system with an unavoidable incomplete elimination of photoresist during fabrication. Detection signals are then observed in the presence of photoresist residues on the P-PAD, even at a blank HRP concentration.

Advances in Microfluidic Paper-Based Analytical Devices for Food and Water Analysis.

Food and water contamination cause safety and health concerns to both animals and humans. Conventional methods for monitoring food and water contamination are often laborious and require highly skilled technicians to perform the measurements, making the quest for developing simpler and cost-effective techniques for rapid monitoring incessant. Since the pioneering works of Whitesides' group from 2007, interest has been strong in the development and application of microfluidic paper-based analytical devices (µPADs) for food and water analysis, which allow easy, rapid and cost-effective point-of-need screening of the targets. This paper reviews recently reported µPADs that incorporate different detection methods such as colorimetric, electrochemical, fluorescence, chemiluminescence, and electrochemiluminescence techniques for food and water analysis.