Extraction and cultivation of pulmonary endothelial cells derived from neonatal mice скачать в хорошем качестве

Extraction and cultivation of pulmonary endothelial cells derived from neonatal mice

Reference: https://app.jove.com/v/2316/isolation... The process of isolating and cultivating pulmonary endothelial cells from neonatal mice is a complex and highly detailed procedure that requires careful attention to various steps to ensure the viability, purity, and functionality of the cells. Initially, the procedure begins with the humane euthanasia of neonatal mice, which is performed in strict accordance with established ethical guidelines and institutional animal care protocols. This step is critical not only for ethical considerations but also to ensure that the tissues obtained are suitable for subsequent isolation processes. Once the mice are euthanized, their lungs are meticulously excised. This requires precision and care to avoid contamination from surrounding tissues or external sources, as any contamination could compromise the integrity of the isolated cells. After the lungs are harvested, the next step involves enzymatic digestion of the lung tissue. This is typically achieved using collagenase or similar enzymes that are specifically chosen for their ability to break down the extracellular matrix components that hold the endothelial cells in place. The enzymatic digestion process is carefully monitored to ensure that the tissue is adequately dissociated without over-digestion, which could lead to cell damage or loss of viability. Once the lung tissue has been digested, the resulting cell suspension is filtered through a mesh or sieve to remove any remaining debris, such as clumps of undigested tissue or dead cells. This filtration step is crucial for obtaining a clean suspension of endothelial cells. Following filtration, centrifugation is performed to concentrate the endothelial cells. During centrifugation, the cell suspension is spun at a specific speed and duration, allowing the denser endothelial cells to pellet at the bottom of the tube while lighter debris remains in the supernatant.