Passage Time - How to Arrange Mammalian Cells and Insect Cells?

Passage Time - How to arrange mammalian cells and insect cells?

1. Cell density Mammalian cells: adherent cultured cells into the logarithmic growth phase, did not meet the state should be submitted when the passage.

Normal cells stop growing when they reach the confluence (contact inhibition) and take longer to recover after re-inoculation.

Transformed cells continue to increase even when confluent but degrade after about two doublings. Similarly, cells cultured in suspension enter the logarithmic growth phase and should also be passaged when they have not reached the state of confluence. When confluence is reached, cells in suspension culture will agglomerate, and the media will become cloudy when the flask is rotated.

Insect cells: insect cells into the logarithmic growth phase, did not meet the confluence should be passed on. Immobilized insect cells can also be passaged when they reach the junction. Although the cells are more likely to dissociate from the incubator at this time, the number of doublings of the cells will decrease if the insect cells are repeatedly passaged at a density exceeding the confluence, Vitality will be reduced, the adherent ability will decline. On the other hand, the passage of the adherent cultured insect cells before confluence requires greater mechanical forces to detach the cells from the monolayer. Repeatedly before reaching the confluence state of reading will lead to reduced cell doubling frequency, decreased vitality, poor health.

2.Media depletion Mammalian cells: Reduced pH in growth media, usually indicative of lactate accumulation, a byproduct of cellular metabolism. Lactic acid is cytotoxic, and decreasing pH is also a negative factor for cell growth. The rate of pH change usually depends on the cell concentration in the culture system. The higher the cell concentration, the faster the medium is depleted. If a rapid decrease in pH is found (> 0.1-0.2 pH units) and the cell concentration is increased, the cells should be passaged.

Insect cells: Growth media for insect cell culture is usually more acidic than mammalian cell culture medium. For example, the pH of TNM-FA and Grace medium for Sf9 cell culture is 6.2. Unlike mammalian cell culture, the pH of the medium in insect cells increases

gradually but does not exceed 6.4. However, like mammalian cells, the pH of growth media begins to decrease as insect cells reach higher densities.

3.passage schedule Strictly according to the scheduled time for cell passage to ensure that the biological behaviour of cells stable, secure to monitor their health status. Gradually adjust cell seeding density from a given seeding density until a steady growth rate, and yield is achieved for the cell. Cells that deviate from the defined growth pattern usually represent poor cell health (e.g., spoilage, contamination) or dysfunction of a component of the culture system (e.g., failure to reach optimal temperature, media too old). We strongly recommend that you keep the detailed cell culture records, record feed and passage times, media used, dissociation methods, seed rating, morphological observations, inoculum concentrations, yields, and antibiotic usage.