The Quality Inspection Of Adult Primary Hepatocytes Culture

ObjectiveWe collected heptocytes from the resected adult benign lesions tissues of liver with the help of surgery to study on quality inspection of adult primary hepatocyte after isolation such as morphological changes,metabolic activities and mycoplasma infection during culture in vitro.MethodsWe isolated hepatocytes from benign liver tissues of patients using modified collagenase perfusion,isolated adult primary hepatocytes counted by blood count plate,liver cells survival rate is checked by trypan blue staining technology.The hepatocyte activity in 7 days continuous culture were detected by CCK-8;.The amount of human Albumin,Urea and Alpha-1 antitrypsin secreted into the medium was measured byenzyme-linked immunosorbent assay(ELISA).Hoechst Staining Method is used for Detection of mycoplasma.Results1.The quantify of fresh adult primary hepatocytes count up to 109,viability detection of fresh adult primary hepatocytes with trypan blue staining was 90%or more.2.The experimental results of CCK-8 showed that the cell viability of hepatocyte were in the best state at the third day of culture.3.Albumin secretion is at its highest concentration at 3rd day and from the 4th day started decreasing gradually.4.Urea secretion during hepatocytes culture was high at 2nd and 3rd day,from 3rd day start decreasing gradually.5.Alpha-1-antitrypsin secretion decreasing gradually during 7days of hepatocytes culture.6.There is no mycoplasma detected in hepatocytes cultured in vitro.Conculsion1.In the process of adult primary liver cell cultivation,Albumin and Urea secretion was high at 3rd day and Alpha-1-antitrypsin secretion decreases gradually during hepatocytes flow culture days.2.There is no mycoplasma infection found during hepatocytes culture so hepatocytes have high purity for transplantation.