Increased sensitivity of NS1 ELISA by heat dissociation in acute dengue 4 cases

Abstract

BackgroundDengue is an acute febrile illness considered the major arboviral disease in terms of morbidity, mortality, economic impact and dissemination worldwide. Brazil accounts for the highest notification rate, with circulation of all four dengue serotypes. The NS1 antigen is a dengue highly conserved specific soluble glycoprotein essential for viral replication and viability that can be detected 0 to 18 days from the onset of fever (peak first 3 days). It induces a strong humoral response and is known as a complement-fixing antigen. Lower NS1 test sensitivity occurs in secondary dengue infections probably due to immune complex formation impairing antigen detection by ELISA.MethodsWe compared the sensitivity of NS1 ELISA in heat dissociated and non-dissociated samples from 156 RT-PCR confirmed acute dengue-4 cases from 362 prospectively enrolled patients.ResultsSecondary infections accounted for 83.3% of cases. NS1 ELISA was positive in 42.5% and indeterminate in 10.2% of dengue-4 cases. After heat dissociation, 7 negative and 16 indeterminate samples turned positive, increasing the overall test sensitivity to 57.7%.ConclusionsAlthough it is time consuming and requires the use of specific laboratory equipment, NS1 ELISA combined with heat dissociation could be a slightly better alternative for triage in suspected dengue cases.

Dengue diagnosis in an endemic area of Peru: Clinical characteristics and positive frequencies by RT-PCR and serology for NS1, IgM, and IgG

“…Although the reported sensitivity of NS1 ELISA was almost four-fold that of IgM ELISA, the value remains lower than those reported in the cited literature. This could be the result of immune complex formation during secondary infection, which impairs NS1 detection by ELISA (Buonora et al, 2017). Some studies have also reported lower NS1 ELISA sensitivity in DENV-4 infections compared with infections caused by other serotypes, in both primary and secondary infections (da Costa et al, 2014).…”

“…The most widely used approach is IgM ELISA because IgM can be detected as early as 4 days after the onset of the disease and in more than 80% of patients with dengue (Rai et al, 2017;Simmons et al, 2012;Teoh et al, 2015). The NS1 antigen can be detected in DENV-infected patients from day 1 up to day 18 after symptom onset and high levels of circulating NS1 correlate with disease severity (Buonora et al, 2017). NS1 can be detected either directly by lateral flow rapid test, also known as immunochromatographic assay (Ambrose et al, 2017), or by ELISA until 9 days later, when RNA detection becomes negative (Sawant et al, 2017).…”

Assessment of diagnostic and analytic performance of the SD Bioline Dengue Duo test for dengue virus (DENV) infections in an endemic area (Savannakhet province, Lao People's Democratic Republic)

“…Previously, immune complex dissociation methods have been described to enhance the sensitivity of antigen detection in dot blot [16] and ELISA applications [10,11,[17][18][19][20][21][22][23]. Nevertheless, the published methods, employing either acid [10,11,16,[19][20][21]23] or heat treatment [11,17,19,22] 3A) were analyzed with the Platelia NS1 Antigen ELISA (Fig 3B), the Panbio μ-capture ELISA (Fig 3C), and the Panbio IgG ELISA (Fig 3D). While a good concordance was observed between the SD Bioline Dengue Duo NS1 test and the Platelia NS1 Antigen ELISA results (Fig 3B and 3E; Cohen's kappa 0.78), only a low percentage of the samples classified as IgM/IgG positive by the Panbio μ-capture and IgG…”

“…Immune complex formation between circulating antigens and antigen-specific antibodies has already been recognized as a factor negatively influencing diagnostic antigen detection in a variety of different disease conditions including HIV infection [20][21][22], West Nile fever [18], and Dengue fever [10,16,17,19,23]. Nevertheless, the DENV IgG assays applied in this work either employ native virus particles displaying all antigens (DENV IgG IIFT) or a recombinant DENV envelope protein E (SD Bioline Dengue Duo IgG rapid test); the antigen used in the Panbio DENV Indirect IgG ELISA has not been specified in the instructions for use.…”

“…Common methods for in vitro immune complex dissociation are acid treatment (using either HCl [20,21] or glycine [10,16,19,21,23], heat exposure [17,19,22], and alkaline treatment [18]. Although all of these methods have been reported to potentially increase accessibility of flavivirus NS1 epitopes in Western Blot and/or ELISA applications [10,16,18,19,23], evidence is strong that these relatively harsh treatments can also prevent efficient antigen detection in some samples.…”