Mouse Anti-VACV L1 Recombinant Antibody (clone 7D11) (CAT#: FAMAB-0715WJ)

Figure 1 Binding to purified VACV.

Serial twofold dilutions of mAb-7D11, 7D11-F(ab')2, and 7D11-Fab or negative control antibody mAb-3D7 starting at 80 μg/ml were tested for the capacity to bind purified virus by VACV-infected-cell lysate ELISA. Some antigen-coated plates were treated with UV light (grey dashed lines) before incubation with mAb-7D11 (MAb-7D11 UV).

Su, H. P., Golden, J. W., Gittis, A. G., Hooper, J. W., & Garboczi, D. N. (2007). Structural basis for the binding of the neutralizing antibody, 7D11, to the poxvirus L1 protein. Virology, 368(2), 331-341.

Figure 2 Binding to recombinant L1.

Serial twofold dilutions of mAb-7D11, 7D11-F(ab')2, and 7D11-Fab or negative control antibody mAb-10F10 (anti-A33) starting at 80 μg/ml were tested for the capacity to bind recombinant L1 by ELISA. Purified L1 at 100 ng/well was untreated or treated with UV light (grey dashed lines). The x-axes in A and B are plotted on log(2) scales.

Su, H. P., Golden, J. W., Gittis, A. G., Hooper, J. W., & Garboczi, D. N. (2007). Structural basis for the binding of the neutralizing antibody, 7D11, to the poxvirus L1 protein. Virology, 368(2), 331-341.

Figure 3 VACV was incubated with purified mAb-7D11, 7D11-F(ab′)2, or 7D11-Fab at 37°C for 1 h.

Virus-antibody mixtures or untreated virus were adsorbed on to confluent monolayers of BSC-1 cells, followed the addition of a semi-solid overlay of 1.5% methylcellulose as described in Methods. After 4 d, plaques were counted after staining with 3% crystal violet. The percent neutralization of plaque formation by antibody addition relative to untreated virus is plotted versus concentration on a log(2) scale.

Su, H. P., Golden, J. W., Gittis, A. G., Hooper, J. W., & Garboczi, D. N. (2007). Structural basis for the binding of the neutralizing antibody, 7D11, to the poxvirus L1 protein. Virology, 368(2), 331-341.

Figure 4 Virus-antibody mixtures or untreated virus were adsorbed on to confluent monolayers of BSC-1 cells, followed the addition of a semi-solid overlay of 1.5% methylcellulose.

After 4 d, plaques were counted after staining with 3% crystal violet. The percent neutralization of plaque formation by antibody addition relative to untreated virus is plotted versus concentration on a log(2) scale.

Su, H. P., Golden, J. W., Gittis, A. G., Hooper, J. W., & Garboczi, D. N. (2007). Structural basis for the binding of the neutralizing antibody, 7D11, to the poxvirus L1 protein. Virology, 368(2), 331-341.

Figure 5 Confluent monolayers of BSC-1 cells were chilled on ice and incubated with ~50 PFU of VACV strain IHD-J per well for 1 h at 4°C.

After adsorption, virus was removed and mAb-7D11 or 7D11-Fab was added to the wells at the indicated concentrations for 1 h at 4°C. After incubation, anti-Fab antibodies (1:100) were added to the wells of some samples for another 1 h at 4°C. Plates were then incubated at 37°C and plaques were visualized as described in A.

Su, H. P., Golden, J. W., Gittis, A. G., Hooper, J. W., & Garboczi, D. N. (2007). Structural basis for the binding of the neutralizing antibody, 7D11, to the poxvirus L1 protein. Virology, 368(2), 331-341.

Figure 6 Interaction of anti-L1 monoclonal antibodies with mutated L1.

COS-7 cells were transfected with pWRG/TPA-L1R (black line), pWRG/TPA-L1R D35N (grey line), or empty pWRG vector (solid grey). Cells were incubated with 1:100 dilutions of mAb-7D11 (anti-L1). A anti-mouse secondary antibody (1:500) conjugated to Alexa 488 fluorochrome was added and cells were analyzed by flow cytometry using a FACSCalibur flow cytometer. For each sample, 10,000 cells were counted.

Su, H. P., Golden, J. W., Gittis, A. G., Hooper, J. W., & Garboczi, D. N. (2007). Structural basis for the binding of the neutralizing antibody, 7D11, to the poxvirus L1 protein. Virology, 368(2), 331-341.

Figure 7 Cell surface expression of modified L1 proteins determined by confocal microscopy.

BS-C-1 cells were transfected with empty vector, pL1op, or psL1op and stained with anti-L1 mAb (7D11) followed by anti-mouse IgG FITC and viewed by confocal microscopy. Upper panel shows confocal fluorescent images and the lower panel shows a merge of confocal fluorescent and differential interference contrast images.

Shinoda, K., Wyatt, L. S., Irvine, K. R., & Moss, B. (2009). Engineering the vaccinia virus L1 protein for increased neutralizing antibody response after DNA immunization. Virology journal, 6(1), 1-8.

Figure 8 Cell surface expression of modified L1 proteins determined by flow cytometry.

BS-C-1 cells were transfected as in panel A. After 24 h, non-permeabilized cells were incubated with MAb 7D11 followed by anti-mouse IgG antibody conjugated to fluorescein isothiocyanate, fixed with paraformaldehyde and analyzed by flow cytometry with gating on L1 positive cells.

Shinoda, K., Wyatt, L. S., Irvine, K. R., & Moss, B. (2009). Engineering the vaccinia virus L1 protein for increased neutralizing antibody response after DNA immunization. Virology journal, 6(1), 1-8.