Recombinant measles proteins, as the improved analogue of whole virus based antigen in ELISA to diagnostics reproach genotypes A and D6
Abstract number: p1664
Kulak M., Netesova N., Belavin P., Ignatyev G.M.
Whole virus based ELISA test systems have different sensitivity to various genotypes. Antigenic variability between different genotypes of measles virus (MV) has existed. We investigated the recombinant proteins NP and HN to develop new antigen with useful properties for applied in ELISA test systems.
Significant antigenic epitopes of nucleoprotein (NP) and haemagglutinin (HN) measles virus strain Edmonston were generated by computer analysis. Using standard gene-engineering techniques was evaluated two fusion peptides NP and HN consist from only linear T-cell antigenic determinants. The virus-neutralization activity of hyperimmune serum on recombinant proteins was determined by plaque reduction neutralization test (PRN). The level of specific IgG in serum to genotypes A, D4, and D6 of measles virus was determined by enzyme-linked immunosorbent assay (ELISA). We used recombinant proteins NP and HN as antigens for ELISA.
Hyperimmune serum was collected from mice after immunization by NP and HN recombinant proteins. The level of neutralize activity was measured in the PRN assay with strain Edmonston. The titre reached up to 1:13.5 and 1:22.9 for NP and HN recombinant proteins, respectively. Interestingly that, hyperimmune serum on recombinant protein NP in ELISA reacted both with NP (titre 1:6400) and with HN (titre 1:3200), and in turn serum on recombinant protein HN reacted only with HN (titre 1:12800). The estimation immunological properties of proteins with use of the panel of serum (50 samples) collected from patients. The diagnosis of measles infection was confirmed in laboratory (by RT-PCR). The nucleotide sequences of RT-PCR products used for genotyping of MV. Selective interaction of antibodies in ELISA with recombinant proteins in relation to various genotypes is revealed. The interaction with genotypes A and D6 was expressed with high level of correlation whereas with genotype D4 any serum did not react authentically (as the control was used recombinant protein N of SARS virus).
We have shown that neutralize antibodies formed hot only on superficial proteins such as HN, F and SH but also on core proteins such as NP. Our data demonstrate that the recombinant proteins NP and HN could be a cost-effective alternative to current whole virus based ELISAs for surveillance for immunity to measles and could more efficient in detecting susceptibility to measles in relation to genotypes A and D6.
|Session name:||XXIst ISTH Congress|
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