METHOD FOR ASSESSING CYTOTOXICITY OF Burcholderia pseudomallei antigens in vitro Russian patent published in 2012 - IPC G01N33/48 C12Q1/00 C12N5/07 

FIELD: medicine.

SUBSTANCE: method provides the use of a cell model for assessing B. pseudomallei antigen toxicity - a melioidosis agent - in vitro. A cytotoxicity reaction is performed on passaged monolayer cell lines of murine fibroblasts L929 or Chinese hamster ovarian cells CHO-K1. Before the experiments, the cells are removed from cold storage; the analysed antigens isolated from various structural components of a bacterial cell of the melioidosis agent are prepared simultaneously at pH 7.0±0.1, sterilised through membrane filters and added in the amount of 20 mcl into wells of primarily prepared 48-well culture plates with a formed monolayer of target cells with 2 wells left with an intact culture (reference). The plate is incubated in a CO₂-incubator at 37°C and atmospheric 5-7% CO₂ saturation for 3 days. The culture plate wells are inspected daily for the presence or the absence of morphological or functional changes in the target cells in comparison with the reference. There shall be observed contact time of a biologically active substance and the cells, concentrations of the substances introduced in the well, morphological and adhesive properties of the cells. The morphological and/or adhesive changes of the cells, including cell death enable stating the presence of cytotoxic action of the analysed antigens on the cells with death of 50% and more cells testifies to manifestations of acute antigen cytotoxicity.

EFFECT: method under the invention is universal, reproducible, applicable for multiple screening of B pseudomallei antigens if working with samples of experimental chemical vaccines, antigen material used for animal immunisation at the stages of producing high-active serums.

2 cl, 5 dwg, 3 ex