Mouse Anti-DNA Recombinant Antibody (clone 1A11) (CAT#: FAMAB-0076CQ)

Figure 1 Binding of purified antibodies to dsDNA in SPR.

The binding of all antibodies was measured using IgG concentrations of 75, 37.5, 18, and 9 nM, as indicated in the legend. Binding of 1A11 (A), 4H8 (B), and 3H9 (C) is displayed in separate panels. Only 3H9 binds dsDNA.

Neeli, I., Richardson, M. M., Khan, S. N., Nicolo, D., Monestier, M., & Radic, M. Z. (2007). Divergent members of a single autoreactive B cell clone retain specificity for apoptotic blebs. Molecular immunology, 44(8), 1914-1921.

Figure 2 L chain mutations include serine 26 to asparagine (S26N) that directs N-linked glycosylation in CDR1 of the 1A11 and 4H8 L chains.

(A) Amino acid sequences for portion of CDR1 are listed for germline, 3H9, 1A11, and 4H8, to illustrate substitutions. (B) Western blot of 4H8, 1A11, and 3H9 incubated in buffer alone (left lane in each set), treated with PNGase F under non-denaturing conditions (center lane), or untreated (right lane). L chains were detected using AP-labeled goat antibodies to mouse kappa. Following PNGase F treatment, mobility of 4H8 and 1A11 L chains increases and differences in migration with 3H9 are eliminated.

Neeli, I., Richardson, M. M., Khan, S. N., Nicolo, D., Monestier, M., & Radic, M. Z. (2007). Divergent members of a single autoreactive B cell clone retain specificity for apoptotic blebs. Molecular immunology, 44(8), 1914-1921.

Figure 3 Binding of 4H8, 1A11, and 3H9 to chromatin is unaffected by PNGase F.

Relative affinities for chromatin in solid phase ELISA were measured, and absorbance readings at 405 nm were plotted. Binding of 3H9 (A), 4H8 (B), and 1A11 (C) is shown in red, along with the binding of PNGase F- treated IgG (green), and with IgG incubated in buffer alone (blue). Triplicate values from two separate experiments were averaged, standard errors were computed, and displayed as vertical bars.

Neeli, I., Richardson, M. M., Khan, S. N., Nicolo, D., Monestier, M., & Radic, M. Z. (2007). Divergent members of a single autoreactive B cell clone retain specificity for apoptotic blebs. Molecular immunology, 44(8), 1914-1921.

Figure 4 Binding of 4H8, 1A11, and 3H9 to apoptotic blebs and nuclear fragments is unaffected by PNGase F.

Jurkat cells, treated with camptothecin to induce apoptosis, were fixed and incubated with 4H8 (A), 1A11 (B), and 3H9 (C). Equivalent reactions were incubated with 4H8 (D), 1A11 (E), and 3H9 (F) treated with PNGase F. Bound antibodies were detected with anti-mouse antibodies (displayed in red). Each of the antibodies, regardless of PNGase F treatment, bound at or near the surface of nuclear fragments. DNA, bound by Sytox Orange, is displayed in blue and annexin V in green. In addition to the composite images, separate wavelength channels are displayed at 1/3 size. Images represent optical cross sections. Controls included cells incubated in the absence of mouse IgG and cells incubated in the presence of z-VAD-fmk, neither of which showed antibody binding.

Neeli, I., Richardson, M. M., Khan, S. N., Nicolo, D., Monestier, M., & Radic, M. Z. (2007). Divergent members of a single autoreactive B cell clone retain specificity for apoptotic blebs. Molecular immunology, 44(8), 1914-1921.