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Information × Registration Number 0224U033298, (0123U102915) , R & D reports Title Features of formation and cryopreservation of spheroids from rat brain cells popup.stage_title Визначення морфологічних особливостей клітин, щовиселяються з сфероїдів, сформованих у краплі протягомрізного часу культивування, в процесі їх культивування напідкладці. Дослідження впливу заморожування-відігріву в присутності кріопротектора ДМСО на морфологічну цілісність сфероїдів та на здатність їх клітин прикріплюватися, мігрувати та розпластуватися в культурі. Визначення оптимального часу еквілібрації сфероїдів з кріопротектором ДМСО перед кріоконсервуванням та оптимального методу відмивання кріопротектора після відігріву кріоконсервованих сфероїдів Head Vsevolodska Svitlana O., Registration Date 24-12-2024 Organization Institute of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine popup.description1 Study of the peculiarities of spheroid formation by brain tissue cells of newborn rats and their subsequent cryopreservation under the protection of penetrating cryoprotectant DMSO. popup.description2 Neural cells (NCs) from the brain of newborn rats are the subject of the study. Aim - To study the characteristics of spheroid (SP) formation by neonatal rat braincells and their subsequent cryopreservation under the protection of the penetrating cryoprotectant DMSO. Research methods - Isolation of cells from neonatal rat brain; monolayer and 3Dcultivation, cryopreservation, fluorescence and light microscopy, morphometry. Experimental studies have demonstrated that freshly obtained NCs from the brain of newborn rats do not form SPs during 3D cultivation by the ‘hanging drop’ method. At the same time, pre-cultured NCs are able to form SPs in a droplet, and the efficiency of spheroid formation depends on the seed concentration and the duration of cultivation. Optimal conditions for the formation of viable SPs formation are concentrations of 12×10 3 , 20×10 3 and 40×10 3 of pre-cultured NPs per 20 µl droplet and a cultivation period of up to 5 days. When transferred to the adhesive surface, the SPs could attach, their cells migrated and formed a monolayer with the morphology of neural and glial cells, indicating high viability of the SPs and the presence of progenitor/stem cells. It was found that only 55±8% of spheroids cryopreserved using the standard two-step protocol, including a 15-minute equilibration step with 10% DMSO at +4°C, retained viability as assessed in monolayer culture. The optimal time for 95% saturation of SP cells with DMSO prior to cryopreservation was found to be 157 seconds at +5°C. The optimal method for removing DMSO from cryopreserved SPs was determined to be a stepwise dilution of the suspension to a DMSO concentration of 0.1%. With this approach, 80% of the spheroids retain their integrity and ability to adhere to the substrate with subsequent migration and differentiation of their cells. Product Description popup.authors Vsevolodska Svitlana O. Kaverinska Hanna I. popup.nrat_date 2024-12-24 Close