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L'Echinococcosi cistica nell'uomo: allestimento di kit da utilizzare nella diagnosi sierologica

Ledda, Salvatore (2012) L'Echinococcosi cistica nell'uomo: allestimento di kit da utilizzare nella diagnosi sierologica. Doctoral Thesis.

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Abstract

Cystic Echinococcosis (CE), caused by E. granulosus, represents a worldwide public health problem. CE serologic diagnosis is useful to confirm clinical infection. The usual source of antigens for immunodiagnostic assays is the hydatid cyst fluid (HCF); Ag 5, a 67 kDa glycoprotein that under reducing conditions dissociate into 38 kDa and 22–24 kDa subunits, is one of the main components of HCF. HCF from naturally infected sheep was analysed by SDS-PAGE, in reducing and non-reducing conditions. Western-Blot (WB) assays, performed against positive human sera, showed a number of protein bands. Positive bands were analysed by mass spectrometry. All of them revealed the presence of Ag5 and/or AgB proteins. HCF was separated by gel filtration (in reducing and in non reducing conditions) and the eluted fractions were loaded on SDS-PAGE. Surprisingly, WB assays revealed the presence of a 60 kDa positive band (possibly Ag5 precursor) into an unreduced gel filtration fraction corresponding to higher molecular weight; positivity was lost by WB in reducing conditions. An ELISA assay was developed using as antigen the purified Ag 5 protein. Sera used for the assay were obtained from 40 patients with clinically diagnosed CE and from 40 uninfected controls. The results showed high specificity (100%) and high sensitivity (95%), indicating its potential as a diagnostic test, particularly in areas where CE is still endemic. Purified Ag 5 may represent a further antigenic marker of infection.

Item Type:Doctoral Thesis
ID Code:6761
Contributors:Cappuccinelli, Pietro
Publisher:Università degli studi di Sassari
Uncontrolled Keywords:Cystic Echinococcosis (CE), ELISA, immunodiagnostic
Subjects:Area 05 - Scienze biologiche > BIO/11 Biologia molecolare
Divisions:001 Università di Sassari > 01 Dipartimenti > Scienze fisiologiche, biochimiche e cellulari
Cicli, scuole e corsi:Ciclo 24 > Scuole > Scienze biomolecolari e biotecnologiche > Microbiologia molecolare e clinica
Deposited On:27 Mar 2012 14:32

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