Sometimes even simple techniques can yield amazing and useful results. A case in point is found in the research of a team from Lyon University in France. The group used gold deposited on electrodes to significantly enhance the on-chip chemiluminescence signal. The result could be a new point-of-care measuring technology for detecting prostate cancer and other diseases that are identified by specific biomarkers.Researcher Christophe A. Marquette, who heads the biochip laboratory at the university, noted that the study was built upon earlier investigations done by the group. The earlier research gave the investigators the basics needed for the recent study — electrode production, gold electrodeposition techniques, biomolecule electroimmobilization and the ability to take sensitive measurements.Researchers built a biochip with enhanced chemiluminescence by printing carbon electrodes on a substrate (a), then depositing gold nanoparticles by forcing a current from the central electrode to the outer ones (b). The gold was functionalized (c) by attaching a biomolecule, which produces a chemiluminescence signal (hν in the diagram). Images reprinted with permission of Langmuir.The researchers began construction of their chemiluminescent signal enhancer by screen-printing carbon electrodes on a substrate. There were eight 0.2-mm2 working electrodes, each beginning at the substrate’s edge and terminating around a ring-shaped pseudoreference electrode, which almost enclosed a central auxiliary electrode. This arrangement allowed them to achieve a three-electrode setup, a key requirement for addressing specific electrodes later in the processing.Next, they electrodeposited gold on each electrode, immersing the carbon electrodes in an acidic solution of hydrogen gold chloride and forcing a current from the central auxiliary to the working electrodes for six minutes. Imaging with a Zeiss optical microscope showed gold nanoparticles dotting the surface. For electrodes in which a –2 μA current was used during electrodeposition, gold coverage was spotty. Those that had undergone a –15 μA current had a uniformly dense gold layer, so the researchers used these electrodes. With an atomic force microscope from NT-MDT Co. of Moscow, they determined that the gold nanoparticles were 800 nm in diameter, with a surface roughness of about 16 nm. Once the gold deposition was completed, the researchers rinsed the electrodes clean and functionalized them, a process that depended critically upon a three-electrode setup. Exploiting that setup, they immobilized biomolecules onto the surfaces of the electrodes. They placed a solution containing the biomolecules of interest on the device and applied a varying voltage to specific electrodes. They functionalized the gold with prostate-specific antigen in one case and a particular oligonucleotide sequence in the other. Rinsing and cleaning the electrodes ensured that all unbound molecules were removed.To test these biochips, they exposed the chips functionalized with prostate-specific antigen to the antiprostate-specific antigen monoclonal antibodies, and they exposed the biochips functionalized with an oligonucleotide sequence to the complementary strand. In both cases, they labeled the target with biotin, which they used to bind to streptavidin labeled with a horseradish peroxidase. The labeling allowed them to get a chemiluminescent burst of light when biomolecular interactions occurred.Using a Fujifilm CCD camera and a 10-min integration time, they measured chemiluminescence from a bare screen-printed carbon electrode and one coated with functionalized gold. They found that the light intensity of the gold-modified screen-printed electrodes was 126 times that of the bare screen-printed electrodes for the DNA target sequence. In the case of antiprostate-specific antigen, the enhancement factor of gold to nongold was 229.Marquette said that the enhancement was not totally unexpected, although it had been thought that the gold nanoparticles might need to be smaller — in the size range of tens of nanometers — to achieve the effect. The fact that the ∋800-nm particles still enhanced the signal was surprising but clarified the mechanism of the enhancement, he said.Using a Kontron spectrophotometer to monitor the 405-nm emission from specially processed electrodes, the researchers showed that the extra surface area of the gold nanoparticles increased the DNA signal twofold. The rest of the increase they attributed to the effect of the gold itself.The enhancement difference between the DNA and prostate-specific antigen, they reported, was the result of quenching — or rather lack of quenching in the case of the monoclonal antiprostate-specific antigen. Such quenching is a function of the inverse fourth power of the distance between the donor (the light-emitting chemical) and acceptor (the gold surface). In the case of the DNA biochips, the distance was 10.2 nm, whereas in the protein biochips, the distance was 25 nm, which explains the difference in enhancement factors.The investigators demonstrated the technology in an immobilized p53 oligonucleotide and prostate-specific antigen antigen assay. They found that they could detect the DNA sequence at a 0.1-nM concentration in a 25-μl sample. They could detect it over two orders of magnitude, from a concentration of 0.1 to 10 nM. With regard to the prostate-specific antigen, the detection limit was 4 ng/ml, a clinically useful result. The work is detailed in the July 31 issue of Langmuir.The researchers plan to investigate whether similar effects can be seen on other electrodeposited metals. Marquette reported also that the group might be looking for an industrial partner to collaborate with for its next research phase.“The next step in our lab is to generate a multiparameter biochip with the developed architecture,” he said.