The possibility of post-vaccination anti-plague immunity evaluation was researched using antigen-stimulated cells tests in vitro and cytometry analysis. The object of study — the blood samples of 17 people immunised by the live plague vaccine (Yersinia pestis EV) epicutaneously. Blood taking was carried out before vaccination and after immunisation on 7 and on 21 days, in 3 and in 6 months. Intensity antigen reactivity of lymphocytes was detected by cell tests in vitro, analysing markers of early (CD45+CD3+CD25+) and late (CD45+CD3+HLA-DR+) lymphocyte activation using flow cytometry. The complex of water-soluble Y. pestis antigens and allergen — pestin PP was tested as antigen. The high stimulating potential was defined of the water-soluble antigens Y. pestis complex. It is shown that coefficient of stimulation of relative level T- lymphocytes which express receptors for IL-2 was positive for all observation times after immunisation. The coefficient of stimulation had maximum values at 21 days (56.37%) and at 3 (47.41%) months. In identifying HLADR-positive lymphocytes before vaccination, the negative coefficient of stimulation was indicated on 7 and 21 days and the positive coefficient of stimulation was indicated at 3 and at 6 months. Analysis of intensity expression of early and late lymphocyte activation markers dynamics showed the possibility and prospect of application of cellular in vitro tests for the laboratory evaluation of specific reactivity of cellular immunity in both the early (7 days) and late (6 months) periods after vaccination. The results can be the basis for developing a new algorithm for assessment of immunological effectiveness of vaccination people against plague. It is the algorithm based on the identification of lymphocyte activation markers by antigen stimulation in conditions in vitro.
Over the years, the production release of the plague vaccine is well developed its technology. The technological cycle of production of the preparation consists of regulated steps, however, despite their effectiveness it is necessary to modernize the manufactoring process, for example, solutions for some of the pressing needs of the customers, in particular, small groups of immunization. Our research has focused on obtaining experimental samples plague vaccine smaller compared to the commercial vaccine, the number of doses per vial prepared in a biomass production unit (ACM-Sh) surface by cultivation using all regulated processing steps, except step of combining content two swabs, and then an additional dilution of the cell suspension stabilizer. However, the time information and the subsequent preparation of such a vaccine is excluded us, since biomass is the second flush in quantitative terms is a ready raw material for the preparation of reduced dosage. The benefits of receiving the vaccine reduced the number of doses directly from the biomass of the second flush with the concentration of microbial cells Yersinia pestis EV 20–40 × 109 biotechnology greatly simplify the manufacture of such a preparation. The experimental vaccine series were tested by major regulated parameters: optical concentration, vitality, thermal stability, the loss on drying. In addition, the vaccine was prefabricated with high baseline viability to extreme temperatures (37±1)°C for 24 hours to exclude enough viable microbial cells for subsequent stabilization indicator of viability during storage. It should be noted that all the experimental samples preserved viability index not lower regulated (25%) during the experiment, in contrast to the commercial preparation. To determine the stability of the formulation during storage (over 3 years) was a comparative analysis of the viability of the experimental and commercial lots. To assess post vaccination immune analyzed the immune response to the introduction of a plague vaccine using FACSCalibur flow cytometer, considering that this technology has a high specificity, sensitivity and informativity. With regard to the immunogenic properties, the active component is recorded at a very high level as the white mice, and guinea pigs. Thus, the main biological indicators derived preparations (viability, thermal stability, storage stability) exceed those of commercial analog and provide effective immunological alterations and highly immunogenic in experimental animals.
Objectiveof the study was to test the nutrient medium based on the enzymatic hydrolysate of corn extract condensed for a scaled production of live plague vaccine and to check the quality of the obtained batches against the specified parameters.Materials and methods.A dense nutrient medium based on corn extract was used to grow biomass in the process of live plague vaccine production. The quality parameters of the vaccine preparation obtained were studied by the regulated methods set forth in the regulatory documentation.Results and conclusions. The vaccine was monitored at all stages of its manufacture, including control of the finished dosage form, in strict accordance with the approved regulatory documentation. All the experimental production series complied with the specified indices. Approbation of the production cycle environment for live plague vaccine manufacturing showed efficiency of the conditions and the possibility of environment’s application in the industrial production of the preparation.
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