FIG. 5.
Myc-tagged gN or gM were transiently expressed in HEK293T cells as described in Materials and Methods. Similarly, wild-type gM was coexpressed with myc-tagged gN (gNmyc) by cotransfection of plasmids encoding each of these HCMV ORFs. The cells were lysed, and myc-tagged proteins were collected with paramagnetic beads conjugated with anti-myc antibodies (Miltenyi Biotec, Auburn, Calif.). After the proteins were washed, they were eluted and separated by SDS-PAGE in 12% gels. The proteins were transferred to a nitrocellulose membrane and probed with (a) an anti-myc MAb (9E10) or (b) affinity-purified human anti-gM/gN antibodies, and antibody binding was detected using an ECL kit (Pierce, Rockford, Ill). The positions of molecular mass standards (in kilodaltons) are indicated to the left of the gel in panel a. The most rapidly migrating bands in panel a represent the dye front of the original gel.