Simultaneous multiple immunoassays in a compact disc-shaped microfluidic device based on centrifugal force.

BACKGROUND: We explored the potential of a microfluidic device based on centrifugal force as an immunoassay platform by examining the imprecision of assays carried out with 200 nL of sample. METHODS: Biotinylated antibodies against alpha-fetoprotein (AFP), interleukin-6 (IL-6), and carcinoembryonic antigen [(CEA); 0.1 g/L each in 15 mmol/L phosphate-buffered saline (PBS) containing 0.1 mL/L Tween 20] were attached to a microcolumn packed with streptavidin-coated particles. A 200-nL sample was then allowed to pass through the microcolumn for 240 s, followed by Alexa 647-labeled detection antibody (7.5 mg/L in 15 mmol/L PBS containing 10 g/L bovine serum albumin). The flow rate was controlled by altering the rotational speed. Up to 104 sandwich type immunoassays were completed within 50 min. RESULTS: For AFP, IL-6, and CEA the detection limits were, respectively, 0.15, 1.25, and 1.31 pmol/L. Inter- and intraassay imprecisions (CVs) were <10% and <20%, respectively, for analyte concentrations >5 pmol/L. The CEA antibody had the lowest affinity according to fluorescence image analysis of the microcolumn region. The result was confirmed in a comparative study using BIAcore 3000. CONCLUSIONS: Day-to-day (total) imprecision (CV) of immunoassays on the compact disc-shaped device are <20%. Analysis of fluorescence images allows rapid ranking of antibodies according to their affinities.

High efficiency electrochemical immuno sensors using 3D comb electrodes.

To realize highly sensitive electrochemical immunoassays, a micro-fabricated three-dimensional (3D) electrode was fabricated and applied to enzyme immuno assay based on production of a redox species. The dimensions of the electrodes are 10 microm in width and 30 microm in height, with 20 microm spacing in between, and the 30 pairs of anode and cathode electrodes made up a single sensor. This structure lead to enhancement of the electrochemical reaction, nearly 100% of trap ratio of redox species. It can be applied to highly sensitive enzyme immuno sensing based on p-aminophenylphosphate (PAPP). Applicability of this technique to the immuno assay for one of the clinical diagnostic marker proteins (alpha-fetoprotein; AFP) from 6 to 500 ng/mL was demonstrated.