Hepatitis B surface antigen assembles in a post-ER, pre-Golgi compartment

Double immunofluorescence of SV24 cells with polyclonal anti-PDI (Fig. 2 a) and 10C3 (Fig. 2 b) shows that they reveal the same reticular pattern which is typical for soluble ER proteins. Similarly, anti-KXsKDEL (Fig. 2 c) and 1D3 (Fig. 2 d) yield identical, reticular staining patterns. This result shows that the distributions of PDI, BiP, and that of the bulk of the other KDEL-proteins which are visualized with anti-KXsKDEL are identical in SV24 cells and together with previous work with these reagents (Tooze et al., 1989; Vaux et al., 1990) validates their use to test the hypothesis that HBsAg assembles in the ER. ER protein which has been suggested to dissolve or to prevent the formation of protein aggregates in the ER (Pelham, 1986) , to help proteins to fold and oligomerize correctly (Gething et al., 1986) , and to prevent incompletely assembled proteins from leaving the ER (Bole et al., 1986) . The 99-kD band probably represents ERp99 (Lewis et al., 1985) , the mouse homologue of 94-kD glucose-regulated protein (G~P94; Mazzarella and Green, 1987) , also known as endoplasmin. Mater and Koch, 1988) . Anti-KX~Zd)EL is a polyclonal rabbit antiserum raised against a mixture of peptides having the general sequence KXXXXXKDEL, where the X positions were synthesized from a mixture of alanine, aspartic acid, histidine, glutamine, lencine, tyrosine, and lysine. It specifically recognizes at least 12 different proteins on the Western blot of an SV24 cell lysate, including ERp99, BiP, PDI, and calreticulin ( Fig. 1) as well as the unidentified lower molecular weight protein recognized by 10C3. A more complete description of the production of these antibodies and further evidence that they