Current serological assays using whole rubella virus (RV) as a target antigen for detecting RV-specific antibodies fail to define specific RV proteins and antigenic determinants such as hemagglutinin (HA) and virus-neutralizing (VN) epitopes of rubella virus. A panel of E1 deletion mutants and a subset of E1-specific monoclonal antibodies (MAb) were used for the initial analysis of HA and VN epitopes of E1 glycoprotein. A peptide region (E1(193) to E1(269)) was found to contain HA and VN epitopes. Using both overlapping synthetic peptides and truncated fusion proteins within this region, the HA epitope defined by MAb 3D9F mapped to amino acid residues E1(214) to E1(240), while two VN epitopes defined by MAb 21B9H and MAb 16A10E mapped to amino acid residues E1(214) to E1(233) and E1(219) to E1(233), respectively. The epitopes defined in this study are recognized by antibody whether or not the epitopes are glycosylated.

Better understanding of cell-mediated immune responses to rubella virus would provide the basis for the development of safe and effective vaccines against rubella and would aid in analysis of the pathophysiology of congenital rubella syndrome. We have expressed individual rubella virus structural proteins, El, E2 and C, via vaccinia virus recombinants. Using the expressed recombinant proteins as antigens, we were able to demonstrate antigen-specific lymphocyte proliferative responses in control individuals and individuals with congenital rubella syndrome. Among the two human groups studied, El glycoprotein proved to be a better immunogen than E2 or C. For the control individuals, significant differences in proliferative responses to the structural proteins El, E2, and C were observed. These differences were not significant in individuals with congenital rubella syndrome. In parallel to the lymphoproliferative responses, immunoglobulin G responses were also found directed mainly to the El glycoprotein. These results suggest that El may be the most important rubella virus antigen to study in determining the domains required for constructing subunit vaccines against rubella.

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The identification of T- and B-cell sites recognized frequently by human populations could provide the basis for selecting the candidate T- and B-cell epitopes for the development of an effective synthetic vaccine against rubella. Rubella virus E1 glycoprotein has been shown to be the predominant antigen to which the majority of human populations develop lymphocyte proliferative and antibody responses. To define the T- and B-cell epitopes of E1 glycoprotein of rubella virus, 23 overlapping synthetic peptides corresponding to more than 90% of the amino acid sequence of E1 were synthesized and tested for their capacities to induce proliferative and antibody responses of 10 seropositive individuals. The most frequently recognized T-cell epitopes were EP19 (residues 324-343), with 7 of 10 responders, and both EP12 (residues 207-226) and EP17 (residues 289-308), with 6 of 10 responders, respectively. Two immunodominant linear B-cell epitopes were mapped to residues 157 to 176 (EP9, 8/10) and 374 to 390 (EP22, 6/10) by using peptide-specific enzyme linked immunosorbent assay.

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