Structural Changes in Insulin at a Soft Electrochemical Interface.

Understanding the interaction of proteins at interfaces, which occurs at or within cell membranes and lipoprotein vesicles, is central to our understanding of protein function. Therefore, new experimental approaches to understand how protein structure is influenced by protein-interface interactions are important. Herein we build on our previous work exploring electrochemistry at the interface between two immiscible electrolyte solutions (ITIES) to investigate changes in protein secondary structure that are modulated by protein-interface interactions.

Ion Transfer Voltammetry with an Electrochemical Pen.

We present a new electrochemical system that combines paper-based sensing and ion-transfer voltammetry, bringing the latter a step closer toward point-of-care applications. Studies at the interface between two immiscible electrolyte solutions (ITIES) are often performed to detect redox-inactive species; unfortunately, due to the inherent instability of the interface, it is rather poorly explored outside specialized laboratories. Here, we address this limitation by combining a pen-like device containing the gelled organic phase with a paper-supported aqueous phase.

Implementation of a plasticized PVC-based cation-selective optode system into a paper-based analytical device for colorimetric sodium detection.

On the example of a colorimetric sodium assay, this work demonstrates the implementation of a classical cation-exchange optode relying on an ionophore-doped plasticized PVC membrane into a paper-based analytical device (PAD). An ion-selective optode (ISO) system has been arranged into a vertically-assembled PAD (vPAD) integrating a pH-buffering function. Capillary force-driven sample liquid transportation through the paper matrix enabled pH-adjustment prior to the optical detection of the analyte cation.

High-Resolution Microfluidic Paper-Based Analytical Devices for Sub-Microliter Sample Analysis.

This work demonstrates the fabrication of microfluidic paper-based analytical devices (µPADs) suitable for the analysis of sub-microliter sample volumes. The wax-printing approach widely used for the patterning of paper substrates has been adapted to obtain high-resolution microfluidic structures patterned in filter paper. This has been achieved by replacing the hot plate heating method conventionally used to melt printed wax features into paper by simple hot lamination.

Fast and single-step immunoassay based on fluorescence quenching within a square glass capillary immobilizing graphene oxide-antibody conjugate and fluorescently labelled antibody.

A single-step, easy-to-use, and fast capillary-type immunoassay device composed of a polyethylene glycol (PEG) coating containing two kinds of antibody-reagents, including an antibody-graphene oxide conjugate and fluorescently labelled antibody, was developed in this study. The working principle involved the spontaneous dissolution of the PEG coating, diffusion of reagents, and subsequent immunoreaction, triggered by the capillary action-mediated introduction of a sample solution. In a sample solution containing the target antigen, two types of antibody reagents form a sandwich-type antigen-antibody complex and fluorescence quenching takes place via fluorescence resonance energy transfer between the labelled fluorescent molecules and graphene oxide.

Distance-Based Tear Lactoferrin Assay on Microfluidic Paper Device Using Interfacial Interactions on Surface-Modified Cellulose.

"Distance-based" detection motifs on microfluidic paper-based analytical devices (μPADs) allow quantitative analysis without using signal readout instruments in a similar manner to classical analogue thermometers. To realize a cost-effective and calibration-free distance-based assay of lactoferrin in human tear fluid on a μPAD not relying on antibodies or enzymes, we investigated the fluidic mobilities of the target protein and Tb(3+) cations used as the fluorescent detection reagent on surface-modified cellulosic filter papers. Chromatographic elution experiments in a tear-like sample matrix containing electrolytes and proteins revealed a collapse of attractive electrostatic interactions between lactoferrin or Tb(3+) and the cellulosic substrate, which was overcome by the modification of the paper surface with the sulfated polysaccharide ι-carrageenan.

Paper-based inkjet-printed microfluidic analytical devices.

Rapid, precise, and reproducible deposition of a broad variety of functional materials, including analytical assay reagents and biomolecules, has made inkjet printing an effective tool for the fabrication of microanalytical devices. A ubiquitous office device as simple as a standard desktop printer with its multiple ink cartridges can be used for this purpose. This Review discusses the combination of inkjet printing technology with paper as a printing substrate for the fabrication of microfluidic paper-based analytical devices (μPADs), which have developed into a fast-growing new field in analytical chemistry.

A single-step enzyme immunoassay capillary sensor composed of functional multilayer coatings for the diagnosis of marker proteins.

A single-step, easy-to-use enzyme immunoassay capillary sensor, composed of functional multilayer coatings, was developed in this study. The coatings were composed of substrate-immobilized hydrophobic coating, hydrogel coating, and soluble coating containing an enzyme-labeled antibody. The response mechanism involved a spontaneous immunoreaction triggered by capillary action-mediated introduction of a sample antigen solution and subsequent separation of unreacted enzyme-labeled antibodies and antigen-enzyme-labeled antibody complexes by the molecular sieving effect of the hydrogel.

Advancements in capillary-assembled microchip (CAs-CHIP) development for multiple analyte sensing and microchip electrophoresis.

This review describes advancements toward the development of a capillary-assembled microchip (CAs-CHIP) for simultaneous multiple analyte sensing and microchip capillary electrophoresis. Development of such an advanced system relies on several factors such as improving the fluid handling technique, creating new biosensing mechanisms, and integrating different functional capillaries into a single CAs-CHIP system. Furthermore, we provide an overview of various functional capillaries that have been established for valving and biosensing applications such as ions, metabolites, proteins, and enzyme activities.

Design of a single-step immunoassay principle based on the combination of an enzyme-labeled antibody release coating and a hydrogel copolymerized with a fluorescent enzyme substrate in a microfluidic capillary device.

A combination of an enzyme-labeled antibody release coating and a novel fluorescent enzyme substrate-copolymerized hydrogel in a microchannel for a single-step, no-wash microfluidic immunoassay is demonstrated. This hydrogel discriminates the free enzyme-conjugated antibody from an antigen-enzyme-conjugated antibody immunocomplex based on the difference in molecular size. A selective and sensitive immunoassay, with 10-1000 ng mL(-1) linear range, is reported.

Capillary-based enzyme-linked immunosorbent assay for highly sensitive detection of thrombin-cleaved osteopontin in plasma.

In this study, a highly sensitive capillary-based enzyme-linked immunosorbent assay (ELISA) has been developed for the analysis of picomolar levels of thrombin-cleaved osteopontin (trOPN), a potential biomarker for ischemic stroke, in human plasma. Using a square capillary coated with 8.5 μg/ml anti-human trOPN capture antibody for ELISA, the linear range obtained was 2 to 16 pM trOPN antigen.

Novel fluorescent probe for highly sensitive bioassay using sequential enzyme-linked immunosorbent assay-capillary isoelectric focusing (ELISA-cIEF).

This paper presents a novel rhodamine diphosphate molecule that allows highly sensitive detection of proteins by employing sequential enzyme-linked immunosorbent assay and capillary isoelectric focusing (ELISA-cIEF). Seven-fold improvement in the immunoassay sensitivity and a 1-2 order of magnitude lower detection limit has been demonstrated by taking advantage of the combination of the enzyme-based signal amplification of ELISA and the concentration of enzyme reaction products by cIEF.

Integration of neuraminidase inhibitor assay into a single-step operation using a combinable poly(dimethylsiloxane) capillary sensor.

The conventional neuraminidase inhibitor assay requiring complicated step-by-step operations was successfully integrated into a "single step" operation using a combinable poly(dimethylsiloxane) (PDMS) capillary (CPC) sensor. In the conventional neuraminidase inhibitor assay, complicated step-by-step operations including initial reaction of an inhibitor and an enzyme, and subsequent reaction with a fluorescent substrate are necessary. Furthermore, optimal pH conditions for the enzymatic reaction differ from those of fluorescence detection.

Bulk- and surface-modified combinable PDMS capillary sensor array as an easy-to-use sensing device with enhanced sensitivity to elevated concentrations of multiple serum sample components.

To enhance sensitivity and facilitate easy sample introduction into a combinable poly(dimethylsiloxane) (PDMS) capillary (CPC) sensor array, PDMS was modified in bulk and on its surface to prepare "black" PDMS coated with a silver layer and self-assembled monolayer (SAM). India ink, a traditional Japanese black ink, was added to the PDMS pre-polymer for bulk modification. The surface was modified by a silver mirror reaction followed by SAM formation using cysteine.

Reagent-release capillary array-isoelectric focusing device as a rapid screening device for IEF condition optimization.

This report describes the fabrication and characterization of a simple and disposable capillary isoelectric focusing (cIEF) device containing a reagent-release capillary (RRC) array and poly(dimethylsiloxane) (PDMS) platform, which allows rapid (within 10 min) screening of cIEF conditions by introducing a sample solution into plural RRCs by capillary action followed by electric field application. To prepare the RRC, covalent immobilization of poly(dimethylacrylamide) (PDMA) was conducted to suppress electro-osmotic flow (EOF), followed by physical adsorption of the mixture of carrier ampholyte (CA), surfactant, labeling reagent (LR), and other additives to the PDMA surface to construct a two-layer structure inside a square glass capillary. When the sample solution containing proteins was introduced into the RRC, physically adsorbed CA, surfactant, and LR can be dissolved and released into the sample solution.

Enzyme-release capillary as a facile enzymatic biosensing part for a capillary-assembled microchip.

A simple capillary enzymatic biosensor was developed. This was prepared by simply coating a dissolvable membrane containing enzyme/s on the inner wall of a square glass capillary. An easy measurement was carried out by capillary force sample introduction with concurrent enzyme release and a reaction with a certain substrate.

Single-drop analysis of various proteases in a cancer cell lysate using a capillary-assembled microchip.

Single-drop analysis of two different real sample solutions (2 microL) while simultaneously monitoring the activity of two sets of ten different proteases on a single microfluidic device is presented. The device, called a capillary-assembled microchip (CAs-CHIP), is fabricated by embedding square glass sensing capillaries (reagent-release capillaries, RRC) in the polydimethylsiloxane (PDMS) lattice microchannel, and used for that purpose. First, the performance reliability was evaluated by measuring the fluorescence response of twenty caspase-3-sensing capillaries on a single CAs-CHIP, and a relative standard deviation of 1.

Current development in microfluidic immunosensing chip.

This review accounts for the current development in microfluidic immunosensing chips. The basic knowledge of immunoassay in relation to its microfluidic material substrate, fluid handling and detection mode are briefly discussed. Here, we mainly focused on the surface modification, antibody immobilization, detection, signal enhancement and multiple analyte sensing.

"Drop-and-Sip" fluid handling technique for the reagent-release capillary (RRC)-based capillary-assembled microchip (CAs-CHIP): sample delivery optimization and reagent release behavior in RRC.

The experimental conditions of the sample delivery inside the reagent-release capillary-based capillary-assembled microchip (RRC-based CAs-CHIP) were optimized and the reagent release procedure in the RRC is discussed. Recently, our group introduced the basic concept of the "drop-and-sip" fluid handling technique (Anal. Chem.

Integration of multianalyte sensing functions on a capillary-assembled microchip: simultaneous determination of ion concentrations and enzymatic activities by a "drop-and-sip" technique.

A general and simple implementation of simultaneous multiparametric sensing in a single microchip is presented by using a capillary-assembled microchip (CAs-CHIP) integrated with the plural different reagent-release capillaries (RRCs), acting as various biochemical sensors. A novel "drop-and-sip" technique of fluid handling is performed with a microliter droplet of a model sample solution containing proteases (trypsin, chymotrypsin, thrombin, elastase) and divalent cations (Ca2+, Zn2+, Mg2+) that passes through the microchannel with the aid of a micropipette as a vacuum pump, concurrently filling each RRC via capillary force. To avert the evaporation of the nanoliter sample volume in each capillary, PDMS oil is dropped on the outlet hole of the CAs-CHIP exploiting the capillary force that results in spontaneous sealing of all the RRCs.