Development of Immune-Chromatographic Monoclonal Test-System for the Detection of Yersinia pseudotuberculosis, Serogroup I

Objective of the study was to develop monoclonal immunoassay for the detection of the pseudotuberculosis agent, serogroup I. Materials and methods. Specific components, that were used for immune-chromatographic test-system development were mouse monoclonal antibodies of hybrid cell lines, obtained to lipopolysaccharide antigen of the outer membrane of the pathogen’s «cold» variant (YP-101N2V4, YP-105S5A10); and rabbit anti-species antibodies against murine immunoglobulins. Particles, (30±2) nm in the diameter, were used to prepare colloidal gold-antibody conjugate. Antibody concentration for conjugation was 10-15 % greater than the D580 exit point on the plateau. For the production of immune-chromatographic test-system a set of membranes - MDI Easypack - manufactured by «Advanced Microdevice», India was deployed. Finished conjugate was applied onto the membrane by means of impregnation. Antibodies in the selected quantities were applied onto the analytical and control membranes via Dispensers. Substrates coated with the conjugate and ready-made working membranes were vacuum dried in a heat cabinet. Assembled immune-chromatographic test-systems were cut off 4.5 mm each and tested for specificity and sensitivity. Results and conclusions. Developed has been immune-chromatographic test-system for the detection of pseudotuberculosis pathogen, serogroup I. Utilized have been monoclonal antibodies of the hybrid cell line YP-105C5A10 in colloidal gold conjugate and monoclonal antibodies of the hybrid cell line YP-101H2B4 in the test line. The test-system allows for the detection of Y. pseudotuberculosis strains, serogroup I, at concentrations varying from 500 ths. m.c.·cm^-3 (8 of the 11 strains under study) up to 4 million m.c.·cm^-3 and does not identify closely related yersinia and heterologous microorganisms in quantities of 100 million m.c.·cm^-3.

возбудитель псевдотуберкулеза серогруппы I, моноклональные антитела, иммунохроматографическая тест-система, pseudotuberculosis agent, serogroup I, monoclonal antibodies, immune-chromatographic test-system

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