Recombinant Mouse Anti-HCMV gB Antibody (7-17) (CAT#: FAMAB-0173CQ)

Figure 1 Recognition of mutant ΔgB651 by AD-1-specific MAb 7-17.

Wild-type (wt) gB and mutant ΔgB651 were expressed in monkey kidney cells infected with recombinant vaccinia viruses. (A) Following labeling with [35S]Met/Cys, infected cell proteins were immune precipitated with the AD-1-specific MAb 7-17, the gB oligomer-specific antibody 27-39, or antibody 58-15, which is directed at an epitope located at the extreme carboxyl terminus of gB, and the precipitated proteins were analyzed by SDS-PAGE under reducing conditions. The migration of the 150-kDa gB precursor protein of gB and the 120-kDa precursor protein of ΔgB651 is indicated in the margin. The migration of the 55-kDa TM cleavage product of gB and the 30-kDa cleavage product of the ΔgB651 mutant are also indicated. Note that ΔgB651 was not precipitated by MAb 58-15. (B) Supernatant from the infected cells was centrifuged at 13,000 × g for 20 min and then precipitated with MAb 7-17. Precipitated proteins were analyzed by SDS-PAGE.

Britt, W. J., Jarvis, M. A., Drummond, D. D., & Mach, M. (2005). Antigenic domain 1 is required for oligomerization of human cytomegalovirus glycoprotein B. Journal of virology, 79(7), 4066-4079.

Figure 2 Oligomerization of ΔgB651.

Monkey kidney cells were infected with a recombinant vaccinia virus expressing ΔgB651 and pulse-labeled with [35S]Met/Cys for 10 min. The cultures were then washed extensively and either harvested immediately (time [T] = 0 min) or incubated in media for 120 min. Infected cell proteins from both cultures were centrifuged through a 5 to 30% sucrose gradient for 16 h. One-milliliter fractions were collected from the bottom of the gradient and precipitated with MAb 7-17. Precipitated proteins were analyzed by SDS-PAGE. Note the increased concentration of precipitated ΔgB651 in fraction 3 in the later chase interval, indicating multimerization. The top of the gradient is indicated.

Britt, W. J., Jarvis, M. A., Drummond, D. D., & Mach, M. (2005). Antigenic domain 1 is required for oligomerization of human cytomegalovirus glycoprotein B. Journal of virology, 79(7), 4066-4079.

Figure 3 Pulse-chase analysis of the ΔgB55.

The mutant ΔgB55 was expressed as a recombinant vaccinia virus, and infected monkey kidney cells were pulse-labeled for 2 min with [35S]Met/Cys and then chased in media for the indicated times. The infected cell proteins were precipitated with either MAb 58-15 or the AD-1-specific MAb 7-17, and the precipitated proteins were analyzed by SDS-PAGE under nonreducing conditions. The migration of the forms of ΔgB55 is indicated in the left margin.

Britt, W. J., Jarvis, M. A., Drummond, D. D., & Mach, M. (2005). Antigenic domain 1 is required for oligomerization of human cytomegalovirus glycoprotein B. Journal of virology, 79(7), 4066-4079.

Figure 4 Analysis of gB cysteine mutants.

Image analysis of gB mutants. gB cysteine mutants were generated and transiently expressed in COS-7 cells grown on glass coverslips. Following fixation in 3% paraformaldehyde, the cells were permeabilized and stained with the AD-1-specific MAb 7-17 or the C-terminal specific MAb 58-15 and developed with FITC-conjugated goat anti-mouse IgG3 or Texas red-conjugated goat anti-mouse IgG2b, respectively. Merged images reveal colocalization of signals resulting in a yellow signal. Wt, wild type.

Britt, W. J., Jarvis, M. A., Drummond, D. D., & Mach, M. (2005). Antigenic domain 1 is required for oligomerization of human cytomegalovirus glycoprotein B. Journal of virology, 79(7), 4066-4079.