Terence G. Henares

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. Some of the clinically important currently implemented on the microfluidic immunoassay chips are C-reactive protein (CRP), prostate specific antigen (PSA), ferritin, vascular endothelial growth factor (VEGF), myoglobin (Myo), cardiac troponin T (cTnT), cardiac troponin I (cTnI), and creatine kinase-cardiac muscle isoform (CK-MB). The emerging microfludic immunosensor technology may be a promising prospect that can propel the improvement of clinical and medical diagnosis.

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. After introducing the fundamentals of μPADs and inkjet printing, it touches on topics such as the microfluidic patterning of paper, tailored arrangement of materials, and functionalities achievable exclusively by the inkjet deposition of analytical assay components, before concluding with an outlook on future perspectives.

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. This patterning technique, in combination with the consideration of device geometry and the influence of cellulose fiber direction in filter paper, led to a model µPAD design with four microfluidic channels that can be filled with as low as 0.5 µL of liquid. Finally, the application to a colorimetric model assay targeting total protein concentrations is shown. Calibration curves for human serum albumin (HSA) were recorded from sub-microliter samples (0.8 µL), with tolerance against ±0.1 µL variations in the applied liquid volume.

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. This concentration range was in the detection window of trOPN antigen in plasma samples. Compared with the conventional microplate-based ELISA, the current capillary technique significantly reduced the amounts of reagent from milliliter to microliter, reduced the analysis time from 8 to 3 h, and had a better sensitivity and detection limit performance from approximately 50 pM down to 2 pM of trOPN antigen. These results indicate that this capillary-based immunoassay is a potential tool for biomarker detection and may be useful in clinical trials and medical diagnostic applications.

Open-type capillary-assembled microchip for rapid, single-step, simultaneous multi-component analysis of serum sample

Here we present an open-type capillary-assembled microchip (CAs-CHIP), demonstrating its ease of fabrication, and use for rapid and simultaneous sensing of different serum components; viz., glucose, cholesterol, and alkaline phosphatase (ALP) and pH. Multiple square glass capillary sensors (outer diameter, 300 μm square) were embedded into a black polydimethylsiloxane (PDMS) microchannel array (300 μm width and depth) generating an open-type CAs-CHIP into which samples were introduced by capillary action. The open-type CAs-CHIP was then embedded into a PDMS reservoir chip where PDMS oil was dropped onto both ends of the capillary to avoid evaporation of the nanoliter sample inside the capillaries. We used fluorescein-based probes to achieve simultaneous multi-component sensing by using a single fluorescence filter and obtained accurate measurements with acceptable day-to-day precision (5.3–8.8%). Discrimination between control and analyte-spiked serum samples was simultaneously accomplished for 4 analytes within 10 min.

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.

Efficient immobilization of the enzyme and substrate for a single-step caspase-3 inhibitor assay using a combinable PDMS capillary sensor array

The caspase-3 inhibitor assay, which is important for drug development was successfully integrated into a “single step” by solving the immobilization problem of enzymes with low activity. Here we used the soluble polyethylene glycol (PEG) coatings for enzyme immobilization to the concave-shaped PDMS surface, and substrate immobilization to the convex-shaped PDMS surface. Then these PDMS structures were combined to form a capillary-structure, which we call a combinable poly (dimethylsiloxane) (PDMS) capillary (CPC) sensor, enabling simultaneous immobilization of two different reactive reagents such as enzymes and fluorescent substrates. The present sensor was disposable, and the fluorescence response was simply obtained by introducing the sample solution by capillary action. The caspase-3 activity was able to be maintained at approximately 90% under −80 °C storage conditions even after 5 months. Importantly, the total reaction time was reduced from an hour to 3–5 min, and simultaneous acquisition of multiple data sets required to determine IC50 value was also successfully achieved using the CPC sensor array.

Inkjet printing of biomolecules for biorecognition

Inkjet printing technology has become prevalent not only for office document printing, but has also gained a lot of attention in academic research and industrial manufacturing. Inkjet printers with the ability to reproducibly deposit known and small volumes of liquids onto specific user selectable spots on a large variety of substrates in a non-contact manner can be regarded as accurate tools for liquid dispensing. Those are highly important in the fields of chemistry, biology, or life sciences.