Isolation And Culture Of Mouse Embryonic Stem Cell-Like

The aim of this paper was to study the isolation, culture and characterization of embryonic stem cell of Kun-ming white mouse.Two methods were used to culture embryo : Method 1: The embryo are incubated to blastula state in TCM199 basal culture medium added 10% FBS under 5%CO 2 and 100% humidity condition at 37℃on the feeder layer made by mouse embryonic fibroblasts (MEF) for 4-6 days.Method 2: The embryo are incubated to blastula state in TCM199 basal culture medium added 10% FBS and 0.1%LIF under the same culture condition without using feeder layer for 1-2 days, the rate of blastula adherence were 43.48% and 31.25% respectively by two methods, and the difference of them was insignificant. The blastula cell attached and proliferated, inner cell mass presented columned growth. The columned inner cell mass were digested many times with 0.125% typsin and 0.02%EDTA, then the cell mass comprised 3-4 cells were separated to culture on feeder layer. the colony of embryonic stem cell were obtained with or without feeder layer system.the colony of embryonic stem cell separated from blastula could gain five strains of embryonic stem cell and go down to posterity F7 in vitro. The stem cell survived and proliferated after anabiosis from freeze preserving state. The colony of stem cell presented cloned growth, the cell edge was slippery and massed compacted bird's nest-like, the limit among cells were misty, the embryonic stem cell were proved by AKP activity staining because of presenting strong masculine...