Enzyme immunoassay with amperometric flow-injection analysis using horseradish peroxidase as a label. Application to the determination of polychlorinated biphenyls
✍ Scribed by Michele Del Carlo; Marco Mascini
- Publisher
- Elsevier Science
- Year
- 1996
- Tongue
- English
- Weight
- 575 KB
- Volume
- 336
- Category
- Article
- ISSN
- 0003-2670
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✦ Synopsis
An amperometric detection system for horseradish peroxidase (HRP) activity was optimized using flow injection analysis (PIA) with glassy carbon as a working electrode. Ferroceneacetic acid was investigated as a co-substrate for the electrochemical detection of HRP. The calculated detection limit for HRP was 2.6x lo-" M with incubation of 30 min. The substrate was used in an electrochemical enzyme immunoassay for polychlorinated biphenyls (PCB). We used a competitive assay, where PCB-protein conjugate (gelatin) was immobilized to the solid phase (microtitre assay plate) and the competition was carried out with PCB standards using a limiting amount of anti-PCB IgG. The extent of the competition was evaluated using a secondary, HRP labelled, IgG; the amount of the enzyme label was detected after 30min of incubation with the substrate. The PCB range was 0.1-50 pgml-'. The overall assay time was 2 h and 30min. The within assay precision, over 6 measurements, was lower than 10% for the entire range.