Isolation And Culture Of Goat Endometiu Stromal Cells

The purposes of the present experiment are to study goat endometrial stromal cell in vitro and to select the optimal condition of the isolation and culture in vitro, also dealt with the morphological modality growth and proliferation characteristic. Different digestion methods and isolation methods were used to prepare the stromal cells. Growth character of goat endometrial stromal cells in vitro were studied through primary culture, The cell's identification, serial subcultivation, determination of growth curve, cell freezed and so on .Through this study , the best density of passage and the preferred way of freezing were derermined. The results as follows:1. The effect of digest with 2g/L collagenase was best at 37℃working 3~4h which cell suspension viscidity was little and gland was large so it was readily to isolate the stromal cell and gland cell. A large amount of the pure stromal cells were able to be separated through 200μm filter—brachytely centrifugalization--sedimentation2. Most stromal cells became adherent after 0.5h of culture. Two shapes of stomal cells were found, spindle and polygonal, The nuclus were egg-shap. Overlapping were found and monolayer were formed after culturing 3~4d. Immunocytochemical staining method showed that this 2 shapes of stromal cells were positive for vimentin, with 95% of positive rate, and negative for cytokeratin.3. When passage, the stromal cell were two days latent period and did not reach platform after cultred 8d with density 1×105/mL subculturing 48 wells culture plate and 0.4mL/well. The cells reached plateform after cultured 6d with 5×105/mL subculturing. The cells could not growth with 5×104 and 1×104 subculturing. The passage cell had evident barred lines when it formed monolayer. The cells volume became bigger after 3 passage.4. This method did not fit for conserving the endometrial stromal cells. The freezing liquid with 10% was profit for Cryopreservation the stromal cell. Serum 20~90%was not obviously difference for Cryopreservation and it was good with 20%serum. The stromal cells on dish bottom was conserved with freezing liquid at -70℃.It was take out and defrosted after three weeks. A lot of cells took off from bottom by microscope.