Article Figures & Data
Figures
- Fig. 1.
Reactivity of a panel of MAbs against serial dilutions of immunoaffinity-purified dengue 2 NS1 in the capture ELISA. NS1 was captured with ELISA plates coated with a 1:1,500 dilution of rabbit anti-NS1 polyclonal antisera and subsequently probed with the MAbs at a dilution of 1:1,000.
- Fig. 2.
Effect of human serum components on the detection of NS1 in the capture ELISA. Immunoaffinity-purified, secreted NS1 was serially diluted in the absence (●) or presence of NHS at dilutions of 1 in 5 (Δ) and 1 in 10 (○) and probed with the type-specific MAb 1H7.4. Data points represent the mean ± standard deviation for four replicates. The dashed line is the mean for the negative control samples (OD490, 0.122 ± 0.012).
- Fig. 3.
Capture ELISA detection of NS1 secreted from Vero cells infected with all four dengue virus serotypes. Type-specific (1H7.4) (A) and group-reactive (3D1.4) (B) MAbs were used as the detection probes for media harvests from DEN1- (♦), DEN2- (●), DEN3- (■), and DEN4 (▴)-infected cells.
Tables
- Table 1.
NS1-specific MAbs used as detection probes in the NS1 capture ELISA
MAba ELISA titerb Linear epitopec Cross-reactivity with other dengue serotypesd DEN1 DEN2 DEN3 DEN4 1H7.4 6.0 + − +++ − − 2C9.4 5.4 + − ++ − − 5H4.4 5.4 + − +++ − − 5H6.3 4.6 − − +++ − + 5B5.3 3.5 − − ++ +++ + 4H3.4 5.3 + +++ +++ +++ +++ 3D1.4 5.2 + +++ +++ +++ +++ 3A5.4 5.7 + +++ +++ +++ +++ ↵a All MAbs used in this study are of the immunoglobulin G1 isotype and were described in detail by Falconar and Young (6).
↵b Titers are expressed as log1050% end points, as measured against purified NS1-coated ELISA plates.
↵c Reactivity of the MAbs with linear epitopes was determined by dot blot analysis against reduced and carboxy-methylated, purified dengue 2 NS1. PEPSCAN analysis has identified the location of some of these epitopes and was reported by Falconar et al. (7). See text for details.
↵d Cross-reactivities were determined by immunoblot analysis of SDS-PAGE-separated SDS lysates of Vero cells infected with the dengue virus serotype prototypes DEN1 (Hawaii), DEN2 (NGC), DEN3 (H87), and DEN4 (H241) and given arbitrary values based on intensity of staining.
- Table 2.
NS1 levels in the sera of dengue 2 virus-infected patients
Type of infectiona Gradeb Serum phase No. positive/no. tested (%) [NS1] (μg/ml)c Primary DF Acute 0/6 (0) —d Convalescent 0/6 (0) — Secondary DF Acute 2/5 (40) 0.6, 3.4 Convalescent 0/5 (0) — Secondary DHF Acute 5/11 (45) 0.07, 0.9, 0.8, 2.4, 15.0 Convalescent 0/11 (0) — ↵a Defined by hemagglutination inhibition titers.
↵b DF, dengue fever; DHF, dengue hemorrhagic fever.
↵c NS1 levels were determined based on a comparison of the mean OD readings of diluted serum samples (three replicates for each dilution) in the ELISA capture assay with a standard curve performed in parallel.
↵d NS1 level is below the detection sensitivity of the assay (4 ng/ml).
