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Automated multiplexed ecl immunoarrays for cancer biomarker proteins

By Karteek Kadimisetty, Spundana Malla, Naimish P. Sardesai, Amit A. Joshi, Ronaldo C. Faria, Norman H. Lee and James F. Rusling

Abstract

Point-of-care diagnostics based on multiplexed protein measurements face challenges of simple, automated, low-cost, and high-throughput operation-with high sensitivity. Herein, we describe an automated, microprocessor-controlled microfluidic immunoarray for simultaneous multiplexed detection of small protein panels in complex samples. A microfluidic sample/reagent delivery cassette was coupled to a 30-microwell detection array to achieve sensitive detection of four prostate cancer biomarker proteins in serum. The proteins are prostate specific antigen (PSA), prostate specific membrane antigen (PSMA), platelet factor-4 (PF-4), and interlukin-6 (IL-6). The six channel system is driven by integrated mitropumps controlled by an inexpensive programmable microprocessor. The reagent delivery cassette and detection array feature channels made by precision-cut 0.8 mm silicone gaskets. Single-wall carbon nanotube forests were grown in printed microwells on a pyrolytic graphite detection chip and decorated with capture antibodies. The detection chip is housed in a machined microfluidic chamber with a steel metal shim counter electrode and Ag/AgCl reference electrode for electrochemiluminescent (ECL) measurements. The preloaded sample/reagent cassette automatically delivers samples, wash buffers, and ECL RuBPY-silica antibody detection nanoparticles sequentially. An onboard microcontroller controls micropumps, and reagent flow to the detection chamber according to a preset program. Detection employs tripropylamine, a sacrificial reductant, while applying 0.95 V vs Ag/AgCl. Resulting ECL light was measured by a CCD camera. Ultra low detection limits of 10-100 fg mL(-1) were achieved in simultaneous detection of the four protein in 36 min assays. Results for the four proteins in prostate cancer patient serum gave excellent correlation with those from single-protein ELISA

Topics: electrogenerated chemiluminescence, ultrasensitive detection, microfluidic devices, prostate-cancer, amplification, diagnostics, arrays, proteome, valves, serum
Publisher: 'American Chemical Society (ACS)'
Year: 2015
DOI identifier: 10.1021/acs.analchem.5b00421
OAI identifier: oai:aran.library.nuigalway.ie/:10379/12133
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