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After digestion with trypsin, alpha-chymotrypsin, or chemical cleavage using CNBr, fragments of the tick-borne encephalitis (TBE) virus glycoprotein were isolated which retained their reactivity with neutralizing monoclonal antibodies defining a denaturation-resistant antigenic domain. Upon immunization of mice, these fragments induced antibodies reactive with the immunizing peptide, the denatured glycoprotein and the native glycoprotein as a constituent of the whole virus. The immune sera revealed the same properties as the monoclonal antibodies that were used to select the fragments for immunization: neutralizing activity; haemagglutination-inhibiting activity; blocking of the binding of antibodies used for selection; enhancement of the binding of other monoclonal antibodies defining a denaturation-sensitive antigenic domain. It was shown that the natural immune response against certain functionally important, denaturation-resistant immunogenic domains on the native protein can be closely mimicked by immunization with defined protein fragments. Antigenic sites present on these fragments may therefore represent essential constituents of a synthetic vaccine. The fine specificities of antibody populations in anti-peptide or anti-protein immune sera were analysed on the basis of single antigenic determinants by blocking assays using radiolabelled monoclonal antibodies that define eight distinct epitopes on the TBE virus glycoprotein. Quantitative differences in the blocking of certain monoclonal antibodies were also observed between human convalescent sera. The establishment of such blocking profiles using a panel of well-characterized monoclonal antibodies may represent a general method for dissecting the specificities of antibody populations present in polyclonal immune sera and could allow investigations on determinant-restricted differences of immune responses and its possible implications for the course of the disease.

Immunogenicity of Tick-borne Encephalitis Virus Glycoprotein Fragments: Epitope-specific Analysis of the Antibody Response