[Comparative evaluation of test-systems for detection of Venezuelan equine encephalomyelitis virus and variola virus using dot immunoenzyme assay and solid-phase lanthanide immunofluorescent assay]. / Sravnitel'naia otsenka test-sistem dlia opredeleniia virusov venesuél'skogo éntsefalomielita loshadei i ospovaktsiny metodami tochechnogo immunofermentnogo analiza i tverdofaznogo lantanidnogo immunofliuorestsentnogo analiza.

The sensitivity and specificity of 4 experimental test systems for dot enzyme immunoassay (dot-EIA) and solid-phase lanthanide immunofluorescent analysis (SP LIFA) were studied with Venezuelan equine encephalomyelitis (VEE) and variolovaccinia viruses. Test systems for SP LIFA proved to be 25 times more sensitive than those for dot-EIA. The test systems were highly specific and did not react with the heterologous viruses and proteins. Diagnostic agent for detecting VEE virus in dot-EIA was false-positive with Sindbis virus in low dilutions. The first trials of the Russian test systems for dot-EIA and SP LIFA showed that these systems rapidly and reliably detect VEE and variolovaccinia viruses in liquid samples.

[Use of 3,3',5,5'-tetramethylbenzidine as substrate in immunoenzyme analysis]. / Ispol'zovanie 3,3',5,5'-tetrametilbenzidina v kachestve substrata pri immunofermentnom analize.

The possibility of using 3,3",5,5'-tetramethylbenzidine and its derivatives in solid-phase enzyme immunoassay and in dot enzyme immunoassay on membrane filters has been demonstrated in experiments with arboviruses. All the tested substrates were fit for specific indication of viral antigens and provided an adequate sensitivity of analyses in comparison with the routinely used substrates, ortho-phenylenediamine and ortho-toluidine. An important advantage of the tested substrates is the absence of carcinogenicity.

[Serodiagnosis and epidemiology of a California encephalitis group of infections in the Ryazan' region]. / Serodiagnostika i épøidemiologiia infektsii gruppy Kaliforniiskogo éntsefalita v Riazanskoi oblasti.

Blood sera of 221 healthy residents of 4 districts of Ryazan region were tested in the neutralization test (NT) with Tahyna and Inkoo viruses. The sum of positive responses in the region was 40.7% on an average, this indicating numerous contacts of the local population with these agents. Blood sera of 520 patients with acute seasonal (May-September) fevers and neuroinfections were tested for antibodies to Tahyna and Inkoo viruses in serologic tests (NT and MAC-ELISA-IgM-antibody capture method). Etiologic relationship of the diseases with Tahyna and Inkoo viruses was detected in 51 (9.8%) cases. The patients developed two types of specific immune response, primary or secondary. The best diagnostic result was attained in testing paired sera of patients in NT and MAC-ELISA. MAC-ELISA helped etiologically diagnose the disease after a single testing of a blood serum collected in the acute period of the disease. Inkoo infection was more incident in subjects aged 21 to 40 in May and July-August. The majority of cases of Tahyna infection were diagnosed in subjects aged under 20 in June and July. The incubation period was from 3 to 7 days, the morbidity was sporadic.

[The rapid detection of arboviruses by dot immunoenzyme analysis on membrane filters]. / Ekspress-indikatsiia arbovirusov metodom tochechnogo immunofermentnogo analiza na membrannykh fil'trakh.

A direct and an indirect variants of dot enzyme immunoassay on membrane filters (dot-EIA) for detection of arbovirus antigens and antibodies were developed. Both variants had similar sensitivity and specificity as compared with the solid-phase enzyme immunoassay usually used for this purpose. The advantages of dot-EIA consist in the possibility of specific detection of antigens in unpurified biological specimens by direct adsorption of the antigen on the membranes, economy aspects, and shortening of the time of analysis.

[The effectiveness of lanthanide immunofluorescence and immunoenzyme analyses in differentiating viruses of the tick-borne encephalitis complex]. / Effektivnost' lantanidnogo immunofliuorestsentnogo i immunofermentnogo analizov pri differentsiatsii virusov kompleksa kleshchevogo éntsefalita.

Comparison of the efficacy of time-resolved fluoroimmunoassay (TR-FIA) and two variants of enzyme immunoassay: conventional (EIA-1) and one using biotin-streptavidin system (EIA-2), in differentiation of viruses of the tick-borne encephalitis complex showed that, their specificity being similar, the TR-FIA method was 2-4 times as sensitive as EIA-2 and 8-32 times as sensitive as EIA-1. When the reactivity of the antigens was assessed by titers, the differentiating capacity was found to be similar and to depend upon the specificity of "sandwich"-forming antibodies and the sensitivity of the immunoassay. When the reactivity of the antigens was assessed by relative reactivity in 4 test systems of different compositions, the differentiating capacity of TR-FIA was higher than that of EIA-1 and EIA-2 which allowed additional differentiation of 2 groups of viruses on the basis of qualitative characteristics of reactivity.

Detection of tick-borne encephalitis virus in ixodid ticks collected in natural foci by time-resolved fluoroimmunoassay.

Time-resolved fluoroimmunoassay (TR-FIA) was used for the first time for evaluation of infestation of ixodid ticks with tick-borne encephalitis virus. Comparison of TR-FIA results with those obtained in enzyme immunoassay and by virus isolation confirmed the high efficacy of the method in question. Positive results of TR-FIA coincided with the data of virus isolation in 83.6% cases, the level of false-negative results did not exceed 1.2%, the overall time consumption amounted to about 1.2 hr.

[The detection of antibodies to the Venezuelan equine encephalomyelitis virus by immunoenzyme analysis using antigen purified in a water-polymer system]. / Vyiavlenie antitel k virusu venesuél'skogo éntsefalomielita loshadei metodom immunofermentnogo analiza s primeneniem ochishchennogo v vodno-polimernoi sisteme antigena.

Human and animal sera were tested for the presence of antibodies to Venezuelan equine encephalomyelitis (VEE) virus by direct enzyme immunoassay (EIA). For the test, the plates were sensitized with a VEE virus preparation purified in a two-phase system of water-soluble polymers. The proposed EIA variant was as specific as that with VEE antigen obtained by fractionation on sucrose cushion, but more sensitive. The high specificity of the assay allowed the antigen purified in the water-polymer system to be used for investigation of antigen relationships among viruses of the VEE complex.

[The differentiation of viruses in the Venezuelan equine encephalomyelitis complex by using monoclonal antibodies and lanthanide immunofluorescence analysis]. / Differentsiatsiia virusov kompleksa venesuél'skogo éntsefalomielita loshadei s primeneniem monoklonal'nykh antitel i lantanidnogo immunofliuorestsentnogo analiza.

Potentialities of differentiation between Venezuelan equine encephalomyelitis (VEE) complex viruses by time-resolved fluoroimmunoassay and enzyme immunoassay were studied. For this, 4 test systems were used based on different combinations of native and labeled polyclonal antibodies to VEE virus, strain Trinidad, and monoclonal (MCA) antibody MAK 14-7 to protein EL of this virus. The maximal sensitivity and specificity was achieved in the test system formed from native MCA MAK 14-7 for sensitization of the solid phase and labeled polyclonal immunoglobulins for demonstration of the test results. This combination of antibodies allowed to differentiate the epidemic variant of VEF/Trinidad (IA) from epizootic variants of Mucambo (III), Pixuna (IV) and attenuated strain No. 230.

Antigenic analysis of tick-borne encephalitis complex viruses by time-resolved fluoroimmunoassay with monoclonal antibodies.

The antigenic structure of 5 strains of tick-borne encephalitis (TBE) virus and 7 other viruses of the TBE complex was examined by the highly sensitive and specific technique of time-resolved fluoroimmunoassay (TR-FIA). A collection of 8 monoclonal antibodies to the Austrian strain. Neudörfl, was used in this study. The findings demonstrate the uniformity of the antigenic structure of TBE viruses from different geographic regions of the USSR. In addition, an epitope was detected which is characteristic of western variants of TBE virus, and another epitope was detected which permits the differentiation of the east-Siberian strain, Aina, from other TBE virus strains. The unique nature of Skalica virus was confirmed, and its similarity, but not identity, to Langat TP-21 virus was shown. Substantial variability in the antigenic structure of some TBE complex viruses was also demonstrated.

[The use of sectional polystyrene plates in the set-up of lanthanide immunofluorescence analysis]. / Ispol'zovanie razbornykh polistirolovykh planshetov pri postanovke lantanidnogo immunofliuorstsentnogo analiza.

The authors have examined the possibility of using sectional polystyrene plates, made in this country, in time-resolved fluoroimmunoassay (tr-FIA) with Venezuelan equine encephalomyelitis and tick-borne encephalitis arboviruses, and with influenza A virus. The plates presensitized with specific antibodies were found fit for the detection of the antigens of the above viruses. These plates are not recommended for the detection of influenza A virus-specific proteins adsorbed directly onto the microplate surface.

[Monoclonal antibodies cross-reacting with the tick-borne encephalitis virus and with the Venezuelan equine encephalomyelitis virus]. / Monoklonal'nye antitela, perekrestno reagiruiushchie s virusom kleshchevogo éntsefalita i virusom venesuél'skogo éntsfalomielita loshadei.

Employment of radioimmunoassay led to the demonstration of serological crossing between tick-borne encephalitis (TBE) virus and Venezuelan equine encephalomyelitis (VEE) virus. Using hybridoma technology, three hybridomas were produced secreting monoclonal antibodies (MAb) cross-reacting with these two viruses. With MAb, the epitope of binding of these antibodies was shown to be located on protein E of TBE virus and protein E1 of VEE virus. Despite the low percentage (14%) of homology of amino acid sequences of these proteins, 12 areas with homology from 24% to 63% were demonstrated. Considering conservative replacements, homology of these areas was 53%-75%. The assumed existence of some of these areas in alpha-helical conformation may explain the observed immunological crossing.

[Dynamics of accumulation of infectious West Nile virus and viral antigens in experimental infection of mice]. / Dinamika nakopleniia infektsionnogo virusa Zapadnogo Nila, virusnykh antigenov pri éksperimental'noi infektsii myshei.

The appearance of the hemagglutinating (HA), complement-fixing (CF) and soluble (S) antigens and infectious virus in the blood and brain of suckling mice inoculated with various doses of West Nile virus was studied. The infectious virus was isolated from the blood and brains of mice as early as 24 hours after inoculation. Its titres increased in parallel in the brain and blood reaching maximum levels by the end of infection. The HA antigen was detected only in the brain at 2-3 days after inoculation. The CF antigen was detected in the blood in a low titre only in the terminal stage of infection; in the brain the CF antigen was detected within 24 hours after infection with a low dose and within 48 hours with a high dose. The S antigen was isolated only from the brain tissue, and in higher titres after infection with a large dose. This may be associated with excess synthesis of the S antigen at a high multiplicity of infection.