title: A simple heat dissociation method increases significantly the ELISA detection sensitivity of the nonstructural-1 glycoprotein in patients infected with DENV type-4.
abstract: The secreted form of the dengue virus (DENV) nonstructural-1 (NS1) glycoprotein has been shown to be useful for the diagnosis of DENV infections in patients' serum samples. In a number of studies, the sensitivity of the commercially available DENV NS1 glycoprotein detection assays was higher against some DENV serotypes (DENV-1>DENV-3>DENV-2=DENV-4) than others and were also lower using patients' serum samples with secondary versus primary DENV infections. In this study, 471 DENV-4 positive acute phase patients' serum samples were selected from a large panel collected in Brazil from March 2011 to October 2012 by RT-PCR and/or virus isolation followed by serotype determination. The sera from primary (n=228) and secondary (n=238) DENV-4 infections were identified using IgM and IgG capture ELISAs. The sensitivity of a commercial DENV NS1 glycoprotein detection ELISA was then assessed when these serum samples were not pre-treated or pre-treated by acid or heat dissociation prior to being tested. Acid and heat dissociation of patients' serum samples with primary and secondary DENV-4 infections increased significantly the sensitivity of the DENV NS1 glycoprotein detection ELISA from 54.4% to 77.2% (p<0.05) and 82% (p<0.05) and from 39.1% to 63.9% (p<0.05) and 73.1% (p<0.05), respectively. Treatment of DENV infected patients' serum samples using simple and rapid heat dissociation step (100°C for 5min) was, therefore, shown to be very useful for increasing the sensitivity of the DENV NS1 glycoprotein detection ELISA using serum samples from either primary or secondary DENV infected patients.
authors.affiliation: Flavivirus Laboratory, Oswaldo Cruz Institute, Rio de Janeiro, Brazil.
authors.forename: Monique da Rocha Queiroz
authors.initials: Mda R
authors.lastname: Lima
authors.affiliation: Flavivirus Laboratory, Oswaldo Cruz Institute, Rio de Janeiro, Brazil.
authors.forename: Rita Maria Ribeiro
authors.initials: RM
authors.lastname: Nogueira
authors.affiliation: Flavivirus Laboratory, Oswaldo Cruz Institute, Rio de Janeiro, Brazil.
authors.forename: Ana Maria Bispo de
authors.initials: AM
authors.lastname: Filippis
authors.affiliation: Flavivirus Laboratory, Oswaldo Cruz Institute, Rio de Janeiro, Brazil.
authors.forename: Priscila Conrado Guerra
authors.initials: PC
authors.lastname: Nunes
authors.affiliation: Flavivirus Laboratory, Oswaldo Cruz Institute, Rio de Janeiro, Brazil.
authors.forename: Carla Santos de
authors.initials: CS
authors.lastname: Sousa
authors.affiliation: Flavivirus Laboratory, Oswaldo Cruz Institute, Rio de Janeiro, Brazil.
authors.forename: Manoela Heringer da
authors.initials: MH
authors.lastname: Silva
authors.affiliation: Flavivirus Laboratory, Oswaldo Cruz Institute, Rio de Janeiro, Brazil. Electronic address: flaviab@ioc.fiocruz.br.
authors.forename: Flavia Barreto dos
authors.initials: FB
authors.lastname: Santos
mesh_terms: D000143:Acids; D000956:Antigens, Viral / Q000097:blood*; D001938:Brazil; D003715:Dengue / Q000175:diagnosis*; D003716:Dengue Virus / Q000276:immunology / Q000302:isolation & purification*; D004797:Enzyme-Linked Immunosorbent Assay; D006023:Glycoproteins / Q000097:blood; D006358:Hot Temperature; D006801:Humans; D012680:Sensitivity and Specificity; D013048:Specimen Handling / Q000379:methods*; D017361:Viral Nonstructural Proteins / Q000097:blood
chemical_list: D000143:Acids; D000956:Antigens, Viral; D006023:Glycoproteins; D017361:Viral Nonstructural Proteins
keywords: DENV-4; Dengue diagnosis; Immune-complex dissociation; NS1 ELISA; secondary infections