ELISA assay was performed with antigens at 0.5µg/ml and antibody at 1µg/ml. ELISA plates were coated on bench overnight in 0.1M Carbonate pH 9.6, 100µl/well, washed once in wash buffer 300µl/well (TBS + 0.1% Tween 20) and blocked for 2 hours in 1% BSA in D-PBS (300µl/well). Antibodies, diluted to working strength in diluent (DPBS + 1% BSA + 0.05% Tween 20 + 0.2% Proclin 950), were added at 100µl/well and incubated for 2 hours (shaken at ambient temperature), followed by 3 x 300µl/well washes. Goat anti Mouse IgG-HRP (Biorad103005) diluted 1 in 2500 in diluent, was added at 100µl/well and incubated with shaking for 1 hour at ambient temperature. Plate was then washed 6 x 300µl/well. TMB (KPL Sureblue 5120-0077) was added at 100µl/well. Reaction for screening assay was stopped by addition of 1M HCl (100µl/well) and absorbance read at 450nm.
Image by Virology Research Services Ltd
Immunofluorescence was carried out in Vero cells. Cells were seeded on coverslips and infected with DENV (serotypes 1, 2, 3 and 4) for 48h at MOI 1. Control coverslips consisted of uninfected Vero cells. After fixation with 4% PFA, samples were stained with Mouse anti-Dengue virus NS1 antibody (AbDENVNS1-DA034 or MAB12136). Antibody was diluted 1:500 and Triton X-100 was used as detergent. Imaging was performed using a Leica SP5 confocal microscope. Immunofluorescence was observed in DENV serotype 1 and DENV serotype 3 infected cells, using AbDENVNS1-DA034 or MAB12136. DENV serotype 1 infected cells (DV1), stained with AbDENVNS1-DA034 are shown above (Virology Research Services Ltd).
MOUSE ANTI-DENGUE VIRUS PAN-SEROTYPE NS1 ANTIBODY (DA034)
Mouse anti Dengue virus NS1 antibody is specific for the NS1 protein of Dengue virus, detecting NS1 from all four Dengue virus serotypes. It demonstrates negligible cross-reactivity with NS1 proteins from Zika virus. A small amount of cross-reactivity has been observed with the NS1 protein from West Nile Virus in a direct ELISA. No cross-reactivity is seen with Chikungunya virus proteins (E1, E2 and C). This antibody is suitable for use in direct ELISA and in sandwich ELISA.
PRODUCT DETAILS – MOUSE ANTI-DENGUE VIRUS PAN-SEROTYPE NS1 ANTIBODY (DA034)
- Mouse anti Dengue virus NS1 IgG1 monoclonal antibody (clone DA034).
- Greater than 95% purity by SDS-PAGE and buffered in PBS, pH7.4.
- Suitable for use in direct ELISA and in sandwich ELISA.
BACKGROUND
The NS1 protein is a major non-structural protein expressed by the Dengue Virus. The NS1 monomer is a glycosylated protein of approximately 45kDa, which associates with lipids and forms a homodimer inside infected cells. It is necessary for viral replication, and is also secreted into the extracellular space as a hexameric lipoprotein particle, which is involved in immune evasion and pathogenesis by interacting with components from both the innate and adaptive immune systems, as well as other host factors. NS1 is one of the major antigenic markers for viral infection with Dengue.
With more than one-third of the world’s population living in areas at risk of transmission, dengue infection is a leading cause of illness and death in the tropics and subtropics (WHO for Guidelines for Diagnosis, Treatment, Prevention & Control, 2009) As many as 100 million people are infected yearly. Dengue is caused by any one of four related viruses transmitted by mosquitoes. There are no vaccines available to prevent infection with dengue virus.
REFERENCES
- Dengue: guidelines for diagnosis, treatment, prevention and control. World Health Organization 2009.
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