Jennifer H. Thomas

An integrated microfluidic biochemical detection system for protein analysis with magnetic bead-based sampling capabilities

This paper presents the development and characterization of an integrated microfluidic biochemical detection system for fast and low-volume immunoassays using magnetic beads, which are used as both immobilization surfaces and bio-molecule carriers. Microfluidic components have been developed and integrated to construct a microfluidic biochemical detection system. Magnetic bead-based immunoassay, as a typical example of biochemical detection and analysis, has been successfully performed on the integrated microfluidic biochemical analysis system that includes a surface-mounted biofilter and electrochemical sensor on a glass microfluidic motherboard. Total time required for an immunoassay was less than 20 min including sample incubation time, and sample volume wasted was less than 50 microl during five repeated assays. Fast and low-volume biochemical analysis has been successfully achieved with the developed biofilter and immunosensor, which is integrated to the microfluidic system. Such a magnetic bead-based biochemical detection system, described in this paper, can be applied to protein analysis systems.

Electrochemical immunoassay moving into the fast lane

The evolution of electrochemical immunoassay has resulted in significant improvements in assay times and detection limits. These developments are traced using a sandwich enzyme immunoassay for IgG as representative. Decreasing the volume of the immunoassay reaction chamber, improving electrochemical detection techniques, and reducing non-specific adsorption have been important factors.

Microdrop analysis of a bead-based immunoassay

The progress to electrochemical detection of a microbead-based immunoassay in small volumes has led to a reduced assay time and lower detection limits. Three electrochemical techniques are described for an immunoassay with detection in a microdrop. The techniques are amperometric detection with a rotating disk electrode (RDE), a microelectrode, and an interdigitated array (IDA) electrode. An enzyme-labeled sandwich immunoassay with mouse IgG as the model analyte is used to demonstrate the three techniques. The microbead assay is carried out in a test tube using a magnet to control bead collection. Once the immunocomplex is formed on the microbead, the beads are transferred to a microdrop where the enzyme, either alkaline phosphatase or β-galactosidase, generates 4-aminophenol (PAP). PAP is oxidized at the electrode with an applied potential of +290 mV vs. Ag/AgCl. For all three techniques, the upper limit of the dynamic range was 1000 ng/ml mouse IgG, and the detection limits were: 50 ng/ml for the RDE, 40 ng/ml for the microelectrode, and 26 ng/ml for the IDA electrode.

Electrochemical immunoassay at a 17β-estradiol self-assembled monolayer electrode using a redox marker

A simple electrochemical immunoassay was demonstrated using a 17beta-estradiol modified electrode. 17beta-estradiol was immobilized on the gold electrode surface with a self-assembly technique. The specific binding between estradiol antibody and 17beta-estradiol on the electrode surface was evaluated by monitoring the change in the electrode response with three hydrophilic redox markers. The decrease in the electrode response for the redox marker was observed, when the antibody was bound to the estradiol self-assembled monolayer (SAM) electrode surface. The change in the electrode response of the redox marker is attributed to the steric hindrance between the antibody on the electrode surface and the redox marker. The relative standard deviation at 30 microg ml(-1) estradiol antibody was 4.1% (n = 3). The competitive reaction between the antigen in the solution and 17beta-estradiol immobilized on the electrode surface for the limited binding sites on the antibody produced an increase in the electrode response with hydroquinone as the marker. The binding affinity of three antigens including 17beta-estradiol to the estradiol antibody was evaluated. Furthermore, the result obtained from this method was compared with the previously reported enzyme binding assay using the biotinylated estradiol and the biotin-immobilized microtiter plate.

An integrated microfluidic biochemical detection system with magnetic bead-based sampling and analysis capabilities

This paper presents the development and characterization of an integrated microfluidic biochemical detection system for fast and low volume immunoassays using magnetic beads, which are used as both immobilization surfaces and bio-molecule carriers. Magnetic bead-based immunoassay, as a typical example of biochemical detection and analysis, has been successfully performed on the integrated microfluidic biochemical analysis system that includes a surface-mounted biofilter and immunosensor on a glass microfluidic motherboard. Total time required for full immunoassay was less than 20 minutes including sample incubation time and sample volume wasted was less than 50 /spl mu/l during five repeated assays. Fast and low volume biochemical analysis has been successfully achieved with the developed biofilter and immunosensor, which is integrated to microfluidic system.

Comb interdigitated arrays (IDA) electrodes for more rapid and sensitive bead-based immunoassay

This research is dedicated to the development of electrode arrays suitable for a more sensitive and faster electrochemical immunoassay system Signal amplification is achieved through coupling the redox cycling of interdigitated array (IDA) electrodes with an enzyme label, /spl beta/-galactosidase. Comb IDAs enhanced the signal 3 times higher than coplanar IDAs. A magnetic bead-based enzyme assay, as a simulation of an immunoassay, has been performed using the comb and coplanar IDAs for detection. Comb IDAs enabled positioning the beads and attached enzyme closer to the sensing electrodes (/spl sim/10 gm). Detection time was less than 1 min. and the limit of detection was 70 amole of /spl beta/-galactosidase.