Method for separating and culturing primary human esophageal epithelial cells

Abstract

The invention provides a method for separating and culturing primary human esophageal epithelial cells. The method comprises the following steps: (1) aseptically taking out 2 cm of a normal oesophagusexcised in the operation of an esophageal cancer patient; (2) longitudinally cutting open the normal oesophagus along the long axis, flattening the cut normal oesophagus, trimming the flattened normal oesophagus to remove subcutaneous connective tissues and muscles, and repeatedly flushing the trimmed normal oesophagus with a pre-cooled PBS buffer solution containing double antibodies; (3) addinga neutral protease to perform digestion; (4) peeling the epithelial layer and the basal layer; (5) cutting epithelial tissues into pieces, and adding collagenase to digest the epithelial tissue pieces; (6) carrying out digestion termination, filtration, centrifugation and washing, re-suspending the obtained material in a conditioned medium, and inoculating a culture bottle with the obtained solution; (7) placing the inoculation bottle in a 37 DEG C and 5% CO2 incubator, and performing culture; (8) carrying out cell passage and purification; and (9) observing and identifying the morphology ofcells. The method for separating and culturing the human esophageal epithelial cells is simple, stable and efficient, and provides a good experimental material for further researches on esophageal mucosal epithelium differentiation, esophageal pathophysiology, toxicology and the cancerogenesis mechanism.

Description

technical field [0001] The invention belongs to the technical field of cell culture of modern biotechnology, and specifically relates to a method for separating and culturing human esophageal epithelial cells. Background technique [0002] Esophageal cancer is one of the most common malignant tumors in my country, with obvious regional characteristics. The death of esophageal cancer accounts for 16.05% of malignant tumors, and the cause of death ranks fourth among malignant tumors after gastric cancer, liver cancer and lung cancer. The pathogenesis of esophageal cancer has always been one of the hot issues of research, but it has not been fully elucidated yet. In recent years, the research on the malignant transformation of normal esophageal epithelial cells caused by various physical and chemical factors, and finally the development of esophageal cancer has gradually received attention, but the first condition for studying the proliferation, differentiation and malignant t...

AI Technical Summary

Problems solved by technology

[0003] At present, there are few normal esophageal epithelial cells available at home and abroad, and there are very few studies on normal esophageal epithelial cell culture methods in vitro, and most of the methods are either unstable, or the steps are complicated and the reagents used are expensive, which seriously restricts the follow-up research work

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