METHOD FOR PRODUCING ORGANOIDS FROM TUMOUR TISSUE OF THE HEAD AND NECK Russian patent published in 2023 - IPC C12N5/09 

FIELD: medicine.

SUBSTANCE: invention relates to biotechnology, cellular biology, and medicine, in particular, to a method for producing three-dimensional cellular organoids from squamous cell tumours of the head and neck. In order to implement the method, the primary material is transported from the operational unit to the cultivation laboratory within a period of no longer than 4 hours while storing the material at +4°C in a transport medium. The tissue is then washed three times in an EDTA solution, then similarly in an antibiotic-antimycotic solution, followed by washing three times from reagents in a buffer solution. The tumour tissue is cleaned from the surrounding peritumoral tissues to the fullest extent. For enzymatic tissue dissociation, recombinant collagenases, or recombinant trypsin, or a combination thereof are used for 60 to 90 minutes at 37°C while constantly stirring on an orbital shaker at a minimum speed of 50 rpm; additionally, recombinant DNAase is used. The volume of the dissociating solution is 5 times greater than the volume of the material. The suspension of cells isolated from the tissue is diluted by 10 times with a buffer solution and passed through a nylon sieve with a cell size of 100 mcm. DMEM/F12 or the 199 medium required to maintain the undifferentiated state of tumour stem cells and prevent epithelial to mesenchymal transition is used as a cultivation medium. 2 v/v% autogenous plasma obtained from the tumour tissue donor is introduced into the cultivation medium. After several days of cultivation, the formed organoids can be extracted for testing or transferred onto a more viscous matrix based on a cultivation medium containing 25 v/v% autogenous plasma.

EFFECT: possibility of producing organoids from tumour tissue of the head and neck.

1 cl, 3 dwg, 1 tbl, 2 ex