Utility of a Commercial Nonstructural Protein 1 Antigen Capture Kit as a Dengue Virus Diagnostic Tool

Abstract

Annually, over 2.5 billion people are at risk for infection with dengue virus (DENV), while between 50 and 100 million people contract the infection. There is an urgent need for alternative diagnostic tools that can detect DENV during acute infection. Recent studies have shown that DENV nonstructural protein 1 (NS1) is detectable in the blood as early as the onset of symptoms and persists well into the convalescent phase of the infection. We evaluated the utility of the Bio-Rad Platelia DENV NS1 antigen capture kit in combination with real-time reverse transcriptase PCR (RT-PCR) and an IgM antibody capture enzyme-linked immunosorbent assay (MAC-ELISA) for refining a new algorithm for the diagnosis of acute-or convalescent-phase DENV infection with a single clinical sample. We tested the Bio-Rad kit with three panels of sera. These panels were designed to evaluate the sensitivities of the NS1 kit for (i) early-convalescent-phase samples, (ii) acute-phase samples with false-negative PCR results, and (iii) IgM-negative convalescent-phase samples from patients with confirmed secondary DENV infections. Results show that NS1 can be detected in 22% of serum samples collected more than 10 days after the onset of illness and in 22% of samples that did not elicit an IgM response. Additionally, NS1 was detected in 37% of the tested acute-phase samples with false-negative PCR results, suggesting that NS1 detection may be valuable in increasing the sensitivity of current acute-phase diagnostics. These results will improve diagnosis with a single acute-phase or early-convalescent-phase sample for disease surveillance and clinical diagnosis.

Detection of Dengue Virus NS1 Antigen in Infected Aedes aegypti Using a Commercially Available Kit

“…The test is a one-step sandwich format enzyme immunoassay for qualitative or semi-quantitative detection of DENV NS1 antigen from all four dengue virus serotypes. 34 Fifty microliters (50 μL) of the supernatant of centrifuged mosquito homogenate suspensions were placed into each well of a 96-well plate and incubated with 150 μL anti-NS1 monoclonal antibody (MAb) conjugated with horseradish peroxidase in phosphate buffer, Tween 20, and fetal calf serum for 90 min at 37 C. When NS1 was present, an immune-complex MAb-NS1-MAb-peroxidase formed and was revealed by adding a chromogenic substrate (tetramethylbenzidine) and H 2 O 2 to initiate color development. The reaction was stopped by the addition of 100 μL of 1 N sulfuric acid.…”

“…Absorbance was determined at 450 nm (A 450 ). Sample A 450 values were compared with those of positive standards included in the kit 34 ; each sample was assayed in triplicate.…”

Acute Febrile Illness Surveillance in a Tertiary Hospital Emergency Department: Comparison of Influenza and Dengue Virus Infections

“…Serum samples were initially tested for DENV by serotype-specific RT-PCR, 18 dengue-specific non-structural protein-1 (NS1), and by DENV-specific immunoglobulin M (IgM) antibody-capture enzyme-linked immunosorbent assay (anti-DENV MAC-ELISA). 19 Serum samples from patients with severe disease (i.e., hospital admission or clinical signs including plasma leakage, fluid accumulation, respiratory distress, bleeding, and organ impairment) and from those who were influenza PCRnegative were subsequently transported on ice to CDC's Bacterial Zoonosis Branch in Atlanta for Leptospira testing. Specimens were screened for IgM antibodies to Leptospira by using the rapid dipstick ELISA ImmunoDOT kit (GenBio, Inc., San Diego, CA).…”

Emerging Pathogens - How Safe is Blood?

“…The diagnostic window period is 3-5 days for IgM antibodies and 2-3 days for NAT [46]. The diagnostic window period for the NS1 antigen test may also be very short [47]. Dengue virus RNA as well as the NS1 antigen are detectable with onset of symptoms [48].…”