Elimination of shrimp endogenous peroxidase background in immunodot blot assays to detect white spot syndrome virus (WSSV)
2003; Inter-Research; Volume: 53; Linguagem: Inglês
10.3354/dao053263
ISSN1616-1580
AutoresWB Zhan, Jianjun Chen, Zuming Zhang, Li Zhou, Hirofumi Fukuda,
Tópico(s)Aquaculture disease management and microbiota
ResumoDAO Diseases of Aquatic Organisms Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsSpecials DAO 53:263-265 (2003) - doi:10.3354/dao053263 Elimination of shrimp endogenous peroxidase background in immunodot blot assays to detect white spot syndrome virus (WSSV) Wen-Bin Zhan1,*, Jing Chen1, Zhi-Dong Zhang1, Li Zhou1, Hideo Fukuda2 1Laboratory of Pathology and Immunology of Aquatic Animals, LMMEC, Ocean University of China, Qingdao 266003, China 2Fish Pathology Laboratory, Tokyo University of Fisheries, Tokyo 108-8477, Japan *Email: wbzhan@mail.ouc.edu.cn ABSTRACT: False positive results were obtained in immunodot blot assays to detect white spot syndrome virus when horseradish peroxidase-conjugated sheep anti-mouse immunoglobin (Ig) serum was used as a secondary antibody with 3-3¹-diaminobenzine tetrahydrochloride dihydrate as the detection substrate. The cause was considered to be a reaction of shrimp endogenous peroxidase (POD) with the substrate. In experiments designed to inhibit POD activity, 9 different reagents were used at different concentrations and for different treatment times. EDTA, sodium azide, HEPES-Na, NaHSO3, H2O2 and phenylthiourea (PTU) were able to inhibit POD activity by 44, 60, 64, 67, 79, and 90%, respectively. Phenylmethylsulfonyl fluoride did not inhibit POD, and neither periodic acid nor H2O2 in methanol were appropriate due to the formation of flocculant precipitates when added to shrimp extracts. It was concluded that of the treatments tested, 10 mM PTU for 2 h yielded optimal inhibition and that such pretreatment of samples eliminates false positive results in immunodot blot assays. KEY WORDS: Shrimp · Peroxidase · WSSV · Immunodot blot Full text in pdf format PreviousNextExport citation RSS - Facebook - Tweet - linkedIn Cited by Published in DAO Vol. 53, No. 3. Online publication date: February 27, 2003 Print ISSN: 0177-5103; Online ISSN: 1616-1580 Copyright © 2003 Inter-Research.
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