Development of Enzyme-Immunoassay for <i>Bacillus anthracis</i> Detection
Objective of the study was to develop enzyme-immunoassay test-kit for the detection of Bacillus anthracis spores. Materials and methods. Microbial cultures from the State Collection of Microorganisms at the premises of Affiliated Branch of the «48th Central Research Institute» of the Ministry of Def...
| Main Authors: | , , , , , , |
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| Format: | Article |
| Language: | Russian |
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Federal Government Health Institution, Russian Research Anti-Plague Institute “Microbe”
2018-10-01
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| Series: | Проблемы особо опасных инфекций |
| Subjects: | |
| Online Access: | https://journal.microbe.ru/jour/article/view/989 |
| _version_ | 1797220786293964800 |
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| author | D. V. Pechenkin O. O. Fomenkov A. V. Eremkin G. D. Elagin G. V. Kuklina G. V. Baramzina S. S. Ipatov |
| author_facet | D. V. Pechenkin O. O. Fomenkov A. V. Eremkin G. D. Elagin G. V. Kuklina G. V. Baramzina S. S. Ipatov |
| author_sort | D. V. Pechenkin |
| collection | DOAJ |
| description | Objective of the study was to develop enzyme-immunoassay test-kit for the detection of Bacillus anthracis spores. Materials and methods. Microbial cultures from the State Collection of Microorganisms at the premises of Affiliated Branch of the «48th Central Research Institute» of the Ministry of Defense of the Russian Federation and BALB/c mice were used in the research. Hybridization of B-lymphocytes with SP2/0-Ag14 myeloma cells was performed according to G. Kohler and C. Milstein procedure in De St. Fazekas and P. Scheidegger modification. Hybridomas were cultured in the peritoneal cavity of BALB/c mice. Ascitic fluids were isolated from mice, precipitated with ammonium sulfate and purified by means of ion-exchange chromatography for preparation of monoclonal antibodies. Specific activity of hybridoma’s supernatants, ascitic fluids, purified monoclonal antibodies was studied by «sandwich» ELISA. Specific components of test-kit were lyophilized in suitable cryoprotective medium. Results and conclusions.We have obtained new hybrid cell lines producing specific monoclonal antibodies against Bacillus anthracisspore antigens and ascitic fluids from which immunoglobulins were isolated. Optimum combinations of monoclonal antibodies as a sensitizer and a component of immunoperoxidase conjugates have been selected. Monoclonal antibodies 272E10G1-272F7A10 provide the highest sensitivity of ELISA for the detection of anthrax microbe spore antigens. Our enzyme-immunoassay test allows for identification of Bacillus anthracis spores in concentrations up to 5,0·105 spores per milliliter. No cross reaction with closely related saprophytes and other heterologous microorganisms in concentrations of 1,0·108 CFU per milliliter is observed. |
| first_indexed | 2024-03-08T13:42:04Z |
| format | Article |
| id | doaj.art-28ce61f268d44d46927640f26172107d |
| institution | Directory Open Access Journal |
| issn | 0370-1069 2658-719X |
| language | Russian |
| last_indexed | 2024-04-24T12:55:04Z |
| publishDate | 2018-10-01 |
| publisher | Federal Government Health Institution, Russian Research Anti-Plague Institute “Microbe” |
| record_format | Article |
| series | Проблемы особо опасных инфекций |
| spelling | doaj.art-28ce61f268d44d46927640f26172107d2024-04-05T16:47:13ZrusFederal Government Health Institution, Russian Research Anti-Plague Institute “Microbe”Проблемы особо опасных инфекций0370-10692658-719X2018-10-0103788210.21055/0370-1069-2018-3-78-82973Development of Enzyme-Immunoassay for <i>Bacillus anthracis</i> DetectionD. V. Pechenkin0O. O. Fomenkov1A. V. Eremkin2G. D. Elagin3G. V. Kuklina4G. V. Baramzina5S. S. Ipatov6Branch of the «48th Central Research Institute» of the Ministry of Defense of the Russian Federation, KirovBranch of the «48th Central Research Institute» of the Ministry of Defense of the Russian Federation, KirovBranch of the «48th Central Research Institute» of the Ministry of Defense of the Russian Federation, KirovBranch of the «48th Central Research Institute» of the Ministry of Defense of the Russian Federation, KirovBranch of the «48th Central Research Institute» of the Ministry of Defense of the Russian Federation, KirovBranch of the «48th Central Research Institute» of the Ministry of Defense of the Russian Federation, KirovBranch of the «48th Central Research Institute» of the Ministry of Defense of the Russian Federation, KirovObjective of the study was to develop enzyme-immunoassay test-kit for the detection of Bacillus anthracis spores. Materials and methods. Microbial cultures from the State Collection of Microorganisms at the premises of Affiliated Branch of the «48th Central Research Institute» of the Ministry of Defense of the Russian Federation and BALB/c mice were used in the research. Hybridization of B-lymphocytes with SP2/0-Ag14 myeloma cells was performed according to G. Kohler and C. Milstein procedure in De St. Fazekas and P. Scheidegger modification. Hybridomas were cultured in the peritoneal cavity of BALB/c mice. Ascitic fluids were isolated from mice, precipitated with ammonium sulfate and purified by means of ion-exchange chromatography for preparation of monoclonal antibodies. Specific activity of hybridoma’s supernatants, ascitic fluids, purified monoclonal antibodies was studied by «sandwich» ELISA. Specific components of test-kit were lyophilized in suitable cryoprotective medium. Results and conclusions.We have obtained new hybrid cell lines producing specific monoclonal antibodies against Bacillus anthracisspore antigens and ascitic fluids from which immunoglobulins were isolated. Optimum combinations of monoclonal antibodies as a sensitizer and a component of immunoperoxidase conjugates have been selected. Monoclonal antibodies 272E10G1-272F7A10 provide the highest sensitivity of ELISA for the detection of anthrax microbe spore antigens. Our enzyme-immunoassay test allows for identification of Bacillus anthracis spores in concentrations up to 5,0·105 spores per milliliter. No cross reaction with closely related saprophytes and other heterologous microorganisms in concentrations of 1,0·108 CFU per milliliter is observed.https://journal.microbe.ru/jour/article/view/989anthraxmonoclonal antibodiesenzyme-linked immunosorbent assay |
| spellingShingle | D. V. Pechenkin O. O. Fomenkov A. V. Eremkin G. D. Elagin G. V. Kuklina G. V. Baramzina S. S. Ipatov Development of Enzyme-Immunoassay for <i>Bacillus anthracis</i> Detection Проблемы особо опасных инфекций anthrax monoclonal antibodies enzyme-linked immunosorbent assay |
| title | Development of Enzyme-Immunoassay for <i>Bacillus anthracis</i> Detection |
| title_full | Development of Enzyme-Immunoassay for <i>Bacillus anthracis</i> Detection |
| title_fullStr | Development of Enzyme-Immunoassay for <i>Bacillus anthracis</i> Detection |
| title_full_unstemmed | Development of Enzyme-Immunoassay for <i>Bacillus anthracis</i> Detection |
| title_short | Development of Enzyme-Immunoassay for <i>Bacillus anthracis</i> Detection |
| title_sort | development of enzyme immunoassay for i bacillus anthracis i detection |
| topic | anthrax monoclonal antibodies enzyme-linked immunosorbent assay |
| url | https://journal.microbe.ru/jour/article/view/989 |
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