How to Passage Cells: Cell Culture Basics Training

Summary

Key Points

• Cell cultures follow a standard growth pattern, requiring passaging during the exponential log phase to maintain healthy cells and maximize experimental yield. 0:21
• Before starting, sterilize the workspace, gather all necessary supplies, and carefully inspect the cell culture for any signs of contamination or deterioration. 0:37
• For adherent cells, remove spent medium, rinse with a calcium and magnesium-free balanced salt solution, then add a gentle cell dissociation reagent like TrypLE Express. 1:04
• Ensure complete cell dissociation by gently tapping the flask and confirming cell release under a microscope, being careful not to overexpose cells to the dissociation reagent. 1:55
• Inactivate the dissociation reagent (e.g., with serum for trypsin or by dilution for TrypLE), transfer cells to a conical tube, and centrifuge to form a well-defined cell pellet. 2:40
• Resuspend the cell pellet in warm complete growth medium, then take a small sample for cell counting using Trypan blue to differentiate between live and dead cells. 3:06
• Based on the cell count, determine the optimal seeding density, add the required fresh medium, and gently pipette the cell solution into new flasks, ensuring even distribution. 3:42
• For suspension cells, take a sample for counting, add fresh medium based on the count, and split the culture into multiple flasks if necessary, ensuring proper capping. 4:48
• Regularly replace the medium for suspension cells every three weeks to eliminate cell debris and metabolic waste, promoting a healthier culture environment. 5:09