Abstract:

BACKGROUND: The methods for primary culture of human chorionic trophoblast cells in vitro are tedious. Moreover, the low cell purity and high cost limit their application in ordinary laboratory.
OBJECTIVE: To establish a simple, convenient and practical method for primary culture of human chorionic trophoblast cells in vitro.
METHODS: Modified trypsinization was used to isolate chorionic trophoblast cells from 5-10 weeks gestational women by 0.0625% trypsin at 37 ℃ for 25-40 minutes. The cells were puvified by differential attachment and digestion method. The inverted microscope was used to observe the shape of cells, and immunocytochemistry was used to identify the source and the purity of cells．
RESULTS AND CONCLUSION: The microscopic examination revealed that following cultures, a large number of round cells freely floated; some cells began to attach 1 hour later, and 70%-80% cells were attached by 24 hours; the number of cells significantly increased by 5-6 days, displaying triangle or polygon shape and patchy spreading growth; the nuclei were large, oval and centralized; some cells were long-fusiform shaped. The 80%-90% confluent cells were passaged by 7-8 days, and completely confluent by 9-10 days. Cell debris did not attached. The passage cells attached 1.5 hours after culture, rapidly grew, and became confluent by 3-4 days. The appearance of each passage cells was similar, displaying epithelioid cell appearance. The 70%-80% cells were  positive for cytokeratin and negative for vimentin. Low density trypsinization, differential attachment and digestion elimination methods are simple and easy to obtain high purity human chorionic trophoblast cells．