Bulletin "Veterinary biotechnology"

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Veterynarna biotekhnolohiia– Veterinary biotechnology, 2023, 42, 81-90 [in Ukrainian]. https://doi.org/10.31073/vet_biotech42-09

SEREDA O.V., e-mail: This email address is being protected from spambots. You need JavaScript enabled to view it., SPYRYDONOV V.G., e-mail: This email address is being protected from spambots. You need JavaScript enabled to view it., TARASOV O.A., e-mail: This email address is being protected from spambots. You need JavaScript enabled to view it., КRYTSIA Y.P., e-mail: This email address is being protected from spambots. You need JavaScript enabled to view it., NYCHYK S.A., e-mail: This email address is being protected from spambots. You need JavaScript enabled to view it.

 

Institute of Veterinary Medicine of the NAAS

 

 

DEVELOPMENTOFANEXPRESSTESTFORTHEDETECTIONOFAFRICANSWINEFEVERVIRUSANTIGENBYTHEMETHODOFMAGNETICIMMUNOFLUORESCENCEANALYSIS

 Introduction. Immuno fluorescence analysis is a diagnostic method that is based on the specific interaction of fluorescent antibodies with homologous antigens. The antigen-antibody complex formed during this interaction is marked with a fluorescent dye, which can be easily detected by fluorescence microscopy or quantitatively assessed by a fluorescence analyzer.

 The goal of the work –production of agarose magnetic microspheres for the development of an express test for the detection of antigens of African swine fever virus (ASFV) in biological secretions by the method of magnetic immunofluorescence analysis.

 Materials and methods. Microspheres were prepared using agarose powder (for molecular biology, Sigma-Aldrich) and magnetite powder (Fe3O4) with particle size <5 μm (Shanghai Knowhow Powder-Tech Co., Ltd.). Fluorescence of the conjugates was measured using the “StepOnePlus Instrument” polymerase chain reaction (PCR) analyzer from Applied Biosystems, USA.

 Results of research and discussion. Immune complexes of magnetic agarose microspheres-ASF-IgY formed with fluorescently labeled antibodies were visualized by fluorescent microscopy. The bright green staining of sepharose particles, incubated with the target antigen ASFV, is visible in the image, while there is no staining of particles incubated with a 1000-fold excess of the control antigen of Classical swine fever virus, which is a visual confirmation of the high sensitivity and specificity of the proposed method.

 Conclusions and prospects for further research. This article describes the development of a magnetic immunofluorescence analysis (MIFA) method using magnetic agarose microspheres (MAM) modified with antibodies. The effectiveness of the modified antibodies on MAM was demonstrated through an express test for the presence of ASFV antigens. The activity of MAM was comparable to that of non-magnetic affinity particles, but they simplified and intensified the process of capturing and isolating target molecules.

 Keywords: immunofluorescence analysis, magnetic agarose microspheres, African swine fever.

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