METHOD FOR OBTAINING PRIMARY MACROPHAGE LINES WITH PRO-INFLAMMATORY PROPERTIES Russian patent published in 2023 - IPC C12N5/786 A61K35/15 

FIELD: biotechnology; regenerative medicine.

SUBSTANCE: invention relates to a method for obtaining a line of macrophages with pro-inflammatory properties. The method for obtaining a line of macrophages with pro-inflammatory properties is characterized by the following: blood is taken into test tubes with EDTA, the blood is diluted with a phosphate buffer solution containing EDTA, the diluted blood is layered on the ficol solution and centrifuged at room temperature, then the buffy coat is taken, washed twice with a phosphate buffer solution containing EDTA, after once with a solution of phosphate buffer containing EDTA and BSA (1%), the resulting cell pellet is incubated with magnetic particles containing antibodies to the CD14 receptor, the cell fraction is passed through a magnetic column and washed off with a solution of phosphate buffer containing EDTA and BSA, and CD14++ monocytes are obtained, which are cultivated in RPMI medium with the addition of bovine serum, L-glutamine, penicillin, streptomycin, macrophage colony-stimulating factor, cells differentiate into macrophages for 7 days, after which gene knockdown is performed, while olig onucleotides used for knockdown are diluted in phosphate buffer, oligonucleotide pairs are annealed, then incubated at room temperature, lipophilic agent Lipofectamine 3000 is used for cell transfection, after adding lipophilic complexes to cells, incubation is carried out for 24 hours, then the medium is changed to RPMI with the addition of bovine serum, L-glutamine, penicillin, streptomycin, 24 hours after the change of medium, phenotype analysis is carried out, as well as analysis of the expression of pro-inflammatory genes and a culture of macrophages with pro-inflammatory properties is obtained, under certain conditions.

EFFECT: obtaining a macrophage cell culture with a stable pro-inflammatory phenotype, suitable for in vivo and in vitro studies.

1 cl, 2 ex