Recombinant Mouse Anti-MNV Antibody (A6.2, 22) (CAT#: PABL-275)

Figure 1 MNV-1 was passaged 5 times through RAW 264.7 cells in the presence of increasing concentrations of the neutralizing MAb A6.2 or a nonneutralizing isotype control antibody. Passage 0 (P0) virus, P3 virus grown with MAb A6.2, and P5 viruses from both conditions were then analyzed in a neutralization assay with 0, 20, 60, and 200 ng MAb A6.2.

Kolawole, A. O., Li, M., Xia, C., Fischer, A. E., Giacobbi, N. S., Rippinger, C. M.,... & Yu, C. (2014). Flexibility in surface-exposed loops in a virus capsid mediates escape from antibody neutralization. Journal of virology, 88(8), 4543-4557.

Figure 2 Mutations in the E′F′ loop of the P domain mediate MNV-1 escape from MAb A6.2 neutralization in culture.

WT MNV and nine A′B′ loop single point mutants (A), nine E′F′ loop single point mutants (B), and three naturally occurring mutants as well as one double mutant (C) were subjected to in vitro MAb A6.2 neutralization. Virus was incubated with indicated concentrations of MAb A6.2 for 30 min at 37°C before infection of RAW 264.7 cells. Viral titers were measured by plaque assay. Percent infectivity of virus titers at different concentrations of MAb A6.2 was calculated relative to the control without MAb A6.2 set to 100%.

Kolawole, A. O., Li, M., Xia, C., Fischer, A. E., Giacobbi, N. S., Rippinger, C. M.,... & Yu, C. (2014). Flexibility in surface-exposed loops in a virus capsid mediates escape from antibody neutralization. Journal of virology, 88(8), 4543-4557.

Figure 3 Mutations in the E′F′ loop of the MNV-1 P domain mediate escape from MAb A6.2 neutralization in mice.

STAT1−/− mice were simultaneously infected with 106 PFU of the indicated recombinant MNV-1 orally and injected with MAb A6.2 (open box) or isotype control MAb (black box) intraperitoneally. Virus titers were measured 48 h postinfection in indicated tissues from animals infected with the wild type (A) or one of the Q298E (B), S299A (C), A382K (D), A382R (E), or L386F (F) mutants with 500 μg MAb, WT-infected animals injected with 15 (G) and 100 (H) μg MAb, and L386F mutant-infected animals injected with 15 (I) and 100 (J) μg MAb, respectively. Data are presented as means ± standard deviations (SD) from at least five mice per condition from at least two independent experiments. Statistical analysis was performed using the t test to compare the virus titer for each tissue between MAb A6.2-injected mice and the isotype control.

Kolawole, A. O., Li, M., Xia, C., Fischer, A. E., Giacobbi, N. S., Rippinger, C. M.,... & Yu, C. (2014). Flexibility in surface-exposed loops in a virus capsid mediates escape from antibody neutralization. Journal of virology, 88(8), 4543-4557.

Figure 4 Binding of recombinant MNV-1 P-domains to MAb A6.2.

ELISA plates were coated overnight at 4°C with bacterially expressed, recombinant MNV-1 P domain A′B′ loop (A) and E′F′ loop (B) mutants at 50 μg/ml in each well. Diluted purified MAb A6.2 followed by secondary antibodies was incubated for 60 min at 37°C. MNV.CR3 P domain was used as a negative control. Data are presented as means ± SE from three independent experiments. The three bars in each set of bars represent three dilutions of purified MAb A6.2: 1, 0.1, and 0.01 μg/ml.

Kolawole, A. O., Li, M., Xia, C., Fischer, A. E., Giacobbi, N. S., Rippinger, C. M.,... & Yu, C. (2014). Flexibility in surface-exposed loops in a virus capsid mediates escape from antibody neutralization. Journal of virology, 88(8), 4543-4557.

Figure 5 Neutralization of MNV by MAb and Fab A6.2.

Equal amounts of MNV were incubated with MAb 6.2, its Fab fragment, or an isotype control antibody and analyzed in a plaque neutralization assay. The assay was repeated twice, and plaque assays were performed in duplicate to calculate standard deviations.

Katpally, U., Wobus, C. E., Dryden, K., Virgin, H. W., & Smith, T. J. (2008). Structure of antibody-neutralized murine norovirus and unexpected differences from viruslike particles.Journal of virology, 82(5), 2079-2088.