The structural protein coding regions of the genomes of Langat virus (strain TP21) and Yelantsev virus, which was originally described to be a low virulence natural isolate of tick-borne encephalitis virus, were cloned and sequenced. These viruses had both been used as experimental live vaccines against tick-borne encephalitis in Czechoslovakia and Russia, respectively. Peptide mapping and monoclonal antibody binding experiments yielded identical reaction patterns for Langat virus and Yelantsev virus which were distinct, however, from the pattern obtained with tick-borne encephalitis virus. Sequence analysis confirmed this distinctiveness and proved that the vaccine strain Yelantsev was also Langat virus. The envelope protein E of both viruses exhibits an 88% amino acid sequence homology with that of tick-borne encephalitis virus. Assessment of the antigenic reactivity and sequence comparison with the E protein of tick-borne encephalitis virus revealed several differences affecting epitopes involved in virus neutralization. These observations suggest that Langat-like virus-based vaccines may not represent the most effective means to achieve protection against tick-borne encephalitis virus.