Conventionally, the electrochemical determination of hydrogen peroxide has been used to selectively measure acetylcholine, choline, glucose and lactate in diverse biological samples. The target analyte is separated using a microbore or cartridge chromatography column. The analyte then passes through an analyte-specific, oxidase enzyme bound to a post-column immobilized enzyme reactor (IMER) that produces the H2O2 which is then detected downstream at the surface of a working electrode in the electrochemical detector's cell.
Platinum electrodes are frequently utilized because they are more selective for H2O2 than standard glassy carbon electrodes. A glassy carbon electrode would have to operate at a much higher applied potential to detect peroxide and would also respond to all of the other electroactive components in the sample. However, platinum requires long equilibration times after cleaning or system changes.
The new wired enzyme electrode (WENZ) offers several advantages vs. the platinum electrode. It exhibits improved sensitivity, better detection limits and improved operational stability. Initial stabilization of the background current is significantly faster than for a conventional platinum electrode. A redox polymer film containing horseradish peroxidase (HRP) is coated on the surface of a glassy carbon electrode. The redox polymer electrically "wires" the peroxidase enzyme to the electrode. It operates at a lower applied potential than a platinum electrode (+ 100 mV vs. Ag/AgCl) and electrochemically reduces hydrogen peroxide generated by a post-column IMER. This enhanced sensitivity is demonstrated in the chromatograms below.
Chromatograms of (a) Ringer's solution and (b) 0.5 nmole lactate using (left) platinum electrode and (right) wired peroxidase electrode.
Chromatograms of 1 nmole glucose (G), 0.23 nmole ascorbic acid (AA) and 0.24 nmole uric acid (UA) on a bare platinum electrode (left). The use of the wired peroxidase electrode (right) effectively increases response.
Redox cycles for an oxidase IMER and wired peroxidase electrode. The peroxidase is covalently linked to a redox hydrogel coated on the surface of a glassy carbon electrode. The oxidized form of the enzyme can be electroreduced through redox centers in the polymer network.
The wired enzyme electrode is offered in kit form. The kit provides a means of preparing and regenerating the electrode for minimal cost. Kits are available in two formats, according to the style of BASi® electrochemical detector being used. Customers already using a BASi® applications kit based on an oxidase IMER will find it very easy to upgrade to the new wired enzyme electrode format.
Chromatograms of 0.5 pmole acetylcholine and choline with (left) platinum electrode and (right) wired peroxidase electrode.
|MF-2096||Reagent refill for either kit|
|CF-8010||Reagent AAO acid washed alumina|
Peroxidase Redox Polymer solution must be kept cold. Kits are shipped via next-day service from West Lafayette, IN, USA. They are packed in special cold-pack containers to preserve enzyme activity. Upon receipt, the kits should be refrigerated (NOT FROZEN) until used.
INTERNATIONAL CUSTOMERS: As noted, the WENZ must be kept cold while in transit and customs. All BASi® WENZ kits are performance tested before shipping. BASi® cannot warrant the performance of these kits if delays or loss of temperature control in shipping or customs occurs. Special shipping instructions must be received before products can be shipped. International couriers that will assure cold delivery are available. This service can be quoted if you do not have your own appropriate shipping company. BASi® will ship the Peroxidase Kits via Federal Express International Priiority. Please request this service be added to your quotation for these kits. Without special shipping instructions from the customer, BASi® will only ship these items on Monday or Tuesday to better allow for time zone shifts, transport logistics and clearance though customs.
UniJet technology is the property of Bioanalytical Systems, Inc. US Patent 5,399,256.
Wired Enzyme sensor technology is licensed from E. Heller and Co. US Patents 5,320,725 and 5,262,035