8 research outputs found
Fabrication of immunosensor microwell arrays from gold compact discs for detection of cancer biomarker proteins
Nanomaterials-Based Electrochemical Immunosensors for Proteins
For this special issue on 90 years of polarography, the following personal account describes how my early research in electrochemistry and polarography in the laboratory of Prof. Petr Zuman led to a major research effort in the determination of proteins for cancer detection and monitoring. It reviews the very recent history of nanoparticle labels and multiplexed detection in protein immunosensors. It then describes our journey of discovery that has led to ultrasensitive protein immunosensors achieved by combining nanostructured electrodes with particles labeled with up to Âœ million enzymes that can detect down to as little as 1 fg mLâ1 protein in diluted serum. Our most mature multiple protein detection system is a microfluidic device with eight sensors coated with 5-nm gold nanoparticles that uses off-line protein detection with heavily labeled magnetic particles. This approach has led to reliable sub pg mLâ1 detection limits for multiple proteins, provides excellent correlation with referee ELISA methods, and is currently being used for validation of panels of biomarkers for oral and prostate cancer. The article ends with a section on future perspectives
Automated Multiplexed ECL Immunoarrays for Cancer Biomarker Proteins
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 micropumps 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-silicaâantibody
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. Ultralow detection limits of 10â100
fg mL<sup>â1</sup> 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