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Individual plant cells’ oxidative bursts observed in real time Huang says the main focus of his lab is individual plant cells by coating a carIn response to signals triggered by infecstudying how chemical messengers such as bon-fiber microelectrode with platinum tion, individual plant cells produce tranneurotransmitters, ROS, and nitric oxide and precisely controlling the platinum’s sient spikes of hydrogen peroxide a few are generated from single cells using elecmilliseconds in duration, Wei-Hua Huang deposition. The detection limit (5 nM trochemical sensors. and colleagues at Wuhan Uni“Our previous work was all versity (China) and the Chinese based on mammalian cells,” he Academy of Agricultural Scisays. “This is the first time we ences report in AC (2009, DOI have collaborated with plant 10.1021/ac901300b). This biologists. Our next research paper is the first to directly goal will focus on the mechamonitor this release from indinisms of the current spikes we vidual plant cells in real time. detected.” Plants produce reactive oxyHong Yang, a chemist spegen species (ROS) such as percializing in the properties of oxide as a rapid defense against nanomaterials at the University pathogens, an innate immune of Rochester, points out that response known as oxidative platinum and Nafion work well burst. The transient oxidative together in other contexts. microbursts that Huang and his Many hydrogen⫺oxygen fuel colleagues have observed resemble the release of neurotrans- A platinum-nanoparticle-coated carbon-fiber microdisk electrode was cells under development use Nafion as a proton exchange mitters by neurons and contrast used to detect peroxide coming from individual oilseed rape cells. membrane and a platinum with the sustained release of percatalyst. oxide previously observed from peroxide) is among the lowest pub“What they have done is quite interestboth bulk leaf tissue and mammalian cells. lished, with a sensor that is small ing,” Yang says. “The novelty here was to “Oxidative burst coming from single enough for single-cell studies. use these materials’ properties on the anaplant cells has not been well-studied,” Previously reported methods for delytical side.” Huang says. “Our results suggest that tecting low amounts of peroxide were Sensitive methods for detecting ROS the mechanisms of production of reacindirect, using an enzyme⫺substrate recould be useful for studying other types tive oxygen species in plant cells and porter. Electrochemical sensors have the of cells as well, says Jesus Aguirre, who animal cells are quite different.” advantage of being able to monitor ROS studies fungal NADPH oxidases at the The production of ROS is regulated production in real time, Huang says. National Autonomous University of by salicylic acid. Huang’s team used Platinum’s electrochemical properties Mexico. benzothiadiazole, a potent synthetic make it ideal for detecting peroxide. Yet ROS produced by NADPH oxidases analog of salicylic acid, to trigger the platinum has a tendency to aggregate dur- are known to regulate sexual developrelease of ROS in individual protoplasts ing deposition, limiting the sensitivity of a ment and growth in fungi, and were from the oilseed rape plant. Protoplasts coated electrode. To get around this tenrecently shown to be important for the are plant cells that have had their cell dency, Huang’s team used Nafion, a suldevelopment of blood cells in Drosowalls either partially or completely refonated tetrafluoroethylene polymer, as a phila. moved enzymatically. “The number of techniques availWhen stimulated with benzothiadia- template for electrodeposition. Complexing the platinum with ammonia (instead able to detect ROS in vivo at the celluzole, the oilseed rape released ROS for of the usual chloride) allowed efficient lar level is limited, and this has hamseveral minutes, but the release came in deposition on the negatively charged pered research in this area,” Aguirre spikes that lasted a few milliseconds. surface. says. “This highly sensitive technique Transgenic plants with an added gene “Nafion has many hydrophilic pores, would allow determining ROS localfor the metabolic enzyme glucose oxiwhich apparently confine the deposition of ization and the activity and kinetics of dase produced an increase in the durathe platinum,” Huang says. “This made it NADPH oxidases under different extion of the wave of spikes and also the possible to prevent the aggregation of perimental conditions.” duration of the individual spikes. —Quinn Eastman platinum particles and have a large effecThe team was able to detect the tiny tive surface area.” amounts of peroxide produced from 10.1021/AC902155S 2009 AMERICAN CHEMICAL SOCIETY
Published on Web 10/05/2009
NOVEMBER 1, 2009 / ANALYTICAL CHEMISTRY
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