Recombinant Human Anti-DENV NS1 Antibody (22NS1) (CAT#: PABL-070)

Figure 1 Flow cytometry profiles for immunoreactivity against NS1 fragments on the yeast surface by three distinct MAbs (4NS1, 16NS1, and 22NS1). pYD1 represents the vector alone, which expresses the Aga2 protein in the absence of NS1.

Flow cytometry. After WNV or DEN-2 infection of BHK21 cells, intracellular NS1 levels were measured by flow cytometry of permeabilized cells as described previously. Two or three days after infection at a multiplicity of infection of 0.1, infected BHK cells were washed three times in PBS, fixed in PBS with 4% paraformaldehyde for 10 min at room temperature, washed twice in PBS, and permeabilized in Hanks ' balanced salt solution containing 10 mM HEPES (pH 7.3), 0.1% saponin, and 0.02% NaN3 (HHSN). For indirect immunofluorescence experiments, cells were resuspended in HHSN and MAb, incubated for 1 h at 4°C, washed three times in HHSN (4°C), resuspended in a 1/250 dilution of fluorescein isothiocyanate-labeled goat anti-mouse IgG (50 μl), and incubated for 1 h on ice in the dark. Cells were subsequently washed three times in HHSN (4°C), fixed in 1% paraformaldehyde in PBS, and stored in the dark prior to analysis on a Becton Dickinson FACSCalibur flow cytometer.

Chung, K.M., Nybakken, G.E., Thompson, B.S., Engle, M.J., Marri, A., Fremont, D.H., Diamond, M.S. (2006). Antibodies against West Nile Virus Nonstructural Protein NS1 Prevent Lethal Infection through Fc Receptor-Dependent and -Independent Mechanisms. JOURNAL OF VIROLOGY, 1340-1351.

Figure 2 In vivo protection of NS1 MAbs against WNV infection.

Six-week-old C57BL/6 mice were inoculated with 102 PFU of WNV subcutaneously and administered a single dose of purified MAbs (500 μg) or ascites fluid (200 μl) at the same time by an intraperitoneal route. The survival curves were constructed using data from one to three independent experiments with 10 animals each.

Chung, K.M., Nybakken, G.E., Thompson, B.S., Engle, M.J., Marri, A., Fremont, D.H., Diamond, M.S. (2006). Antibodies against West Nile Virus Nonstructural Protein NS1 Prevent Lethal Infection through Fc Receptor-Dependent and -Independent Mechanisms. JOURNAL OF VIROLOGY, 1340-1351.

Figure 3 A model of the topography of WNV NS1 and antibody binding sites.

Nine, one (8NS1), one (22NS1), and eight of our MAbs recognized amino acids 1 to 124, amino acids 1 to 157, amino acids 158 to 235, and amino acids 236 to 352, respectively. 10NS1 recognized both FR-I and FR-II. 17NS1 recognized both FR-I and FR-III. FR-I, FR-II, and FR-III indicate amino acids 1 to 157, amino acids 158 to 235, and amino acids 236 to 352, respectively. Numbers indicate the amino acid position on WNV NS1.

Chung, K.M., Nybakken, G.E., Thompson, B.S., Engle, M.J., Marri, A., Fremont, D.H., Diamond, M.S. (2006). Antibodies against West Nile Virus Nonstructural Protein NS1 Prevent Lethal Infection through Fc Receptor-Dependent and -Independent Mechanisms. JOURNAL OF VIROLOGY, 1340-1351.