METHOD FOR PRODUCING CELL CULTURE OF RETINAL PIGMENT EPITHELIUM FROM ADULT CADAVER DONOR'S EYE

摘要

FIELD: medicine.;SUBSTANCE: producing retinal pigment epithelium (RPE) cell culture from adult cadaver donor's is ensured by removing a corneal-scleral disk from cadaver donor's eyeball. If all parts of choroid occurs to be continued, the eyeball free from the corneal-scleral disk is completely immersed into a container with phosphate-saline buffer with pH 7.4; the sclera is completely separated from the choroid by separating the sclera by meridian incision 2, 3 or 4 petals back and transecting from a suprachoroid space of vortex veins and an intrascleral portion of a visual nerve; that is followed by making three incisions of a choroid-pigment complex (CPC) consisting of the choroid and RPE: first circular incision at 1 mm from an ora serrata, one meridian incision in any meridian in the direction from the first circular incision backwards to an optic disk stump, and a second circular incision at 1 mm from the optic disk stump. Further, the choroid is carefully separated from a neural retina by means of forceps and placed on the bottom of a sterile Petri dish in a position of PRE cell upwards, filled with 3 ml of a mixture of 0.25% trypsin and Versene solution in ratio 1:1 by volume, incubated at 37°C in the CO₂ concentration of 5% for 25 minutes; thereafter, the RPE is separated from the choroid surface by means of phosphate-saline buffer jet with pH 7.4. The produced cell suspension is collected into a test tube, centrifuged; supernatant is poured off; the produced deposit is added with a nutrient medium of the following composition: Dulbecco modified Eagle's medium with Ham's medium F12 - 89%, foetal calf serum - 10%, mixed antibiotics, including penicillin 10000 International Units/ml, streptomycin 10000 mcg/ml, amphotericin 25 mcg/ml - 1%. The cell culture is incubated at 37°C in the CO₂ concentration of 5%; fresh nutrient medium is added every 3-4 days.;EFFECT: invention enables reducing a degree of cell culture contamination.;2 dwg, 1 ex