A simple method to produce 2D and 3D microfluidic paper-based analytical devices for clinical analysis.

This paper describes the fabrication of 2D and 3D microfluidic paper-based analytical devices (µPADs) for monitoring glucose, total protein, and nitrite in blood serum and artificial urine. A new method of cutting and sealing filter paper to construct µPADs was demonstrated. Using an inexpensive home cutter printer soft cellulose-based filter paper was easily and precisely cut to produce pattern hydrophilic microchannels. 2D and 3D µPADs were designed with three detection zones each for the colorimetric detection of the analytes. A small volume of samples was added to the µPADs, which was photographed after 15 min using a digital camera. Both µPADs presented an excellent analytical performance for all analytes. The 2D device was applied in artificial urine samples and reached limits of detection (LODs) of 0.54 mM, 5.19 µM, and 2.34 µM for glucose, protein, and nitrite, respectively. The corresponding LODs of the 3D device applied for detecting the same analytes in artificial blood serum were 0.44 mM, 1.26 µM, and 4.35 µM.

Elucidation of carbohydrate molecular interaction mechanism of recombinant and native ArtinM.

The quartz crystal microbalance (QCM) technique has been applied for monitoring the biorecognition of ArtinM lectins at low horseradish peroxidase glycoprotein (HRP) concentrations, using a simple kinetic model based on Langmuir isotherm in previous work.18 The latter approach was consistent with the data at dilute conditions but it fails to explain the small differences existing in the jArtinM and rArtinM due to ligand binding concentration limit. Here we extend this analysis to differentiate sugar-binding event of recombinant (rArtinM) and native (jArtinM) ArtinM lectins beyond dilute conditions. Equivalently, functionalized quartz crystal microbalance with dissipation monitoring (QCM-D) was used as real-time label-free technique but structural-dependent kinetic features of the interaction were detailed by using combined analysis of mass and dissipation factor variation. The stated kinetic model not only was able to predict the diluted conditions but also allowed to differentiate ArtinM avidities. For instance, it was found that rArtinM avidity is higher than jArtinM avidity whereas their conformational flexibility is lower. Additionally, it was possible to monitor the hydration shell of the binding complex with ArtinM lectins under dynamic conditions. Such information is key in understanding and differentiating protein binding avidity, biological functionality, and kinetics.

Jacalin interaction with human immunoglobulin A1 and bovine immunoglobulin G1: affinity constant determined by piezoelectric biosensoring.

The affinity of the D-galactose-binding lectin from Artocarpus heterophyllus lectin, known as jacalin, with immonuglobulins (Igs) was determined by biofunctionalization of a piezoelectric transducer. This piezoelectric biofunctionalized transducer was used as a mass-sensitive analytical tool, allowing the real-time binding analysis of jacalin-human immunoglobulin A1 (IgA(1)) and jacalin-bovine IgG(1) interactions from which the apparent affinity constant was calculated. The strategy was centered in immobilizing jacalin on the gold electrode's surface of the piezoelectric crystal resonator using appropriate procedures based on self-assembling of 11-mercaptoundecanoic acid and 2-mercaptoethanol thiol's mixture, a particular immobilization strategy by which it was possible to avoid cross-interaction between the proteins over electrode's surface. The apparent affinity constants obtained between jacalin-human IgA(1) and jacalin-bovine IgG(1) differed by 1 order of magnitude [(8.0 ± 0.9) 10(5) vs (8.3 ± 0.1) 10(6) L mol(-1)]. On the other hand, the difference found between human IgA(1) and human IgA(2) interaction with jacalin, eight times higher for IgA(1), was attributed to the presence of O-linked glycans in the IgA(1) hinge region, which is absent in IgA(2). Specific interaction of jacalin with O-glycans, proved to be present in the human IgA(1) and hypothetically present in bovine IgG(1) structures, is discussed as responsible for the obtained affinity values.