A series of cell cultures was performed at three different cell concentrations and cells were counted daily until plateau. Source: Animal Cell Culture: A Guide to Basic Techniques, Fifth Edition
Operation method
Scheme 21.9 Plotting of growth curves of cells in suspension culture
Principle
Series of cell cultures were performed at three different cell concentrations and cells were counted daily until plateau. Move makings For more product details, please visit Aladdin Scientific website.
non-sterile
Suspension cell culture
Culture medium, l00 ml
D-PBSA (for cell counting)
24-well plate
Non-sterile
Plastic box for plates
CO2 incubator or 5% C02 to fill plastic box
Procedure
1. Add the cell suspension from the growth solution to the wells at different concentrations as in monolayer culture (see protocol 21.8).
2 . Sample 0.4 ml from each of the three wells at different times, taking care that the cells are well mixed and completely dispersed as singles.
3. Count with an electronic cell counter (see section 21.1.2) or with a blood cell counter (see section 21.1.1).
4 . Calculate the cell concentration for each sample, plotting linearly on a logarithmic scale with time as for monolayer cell growth (see Section 21.9.3). Cell densities are not suitable for suspension cultures as they do not adhere.
Tip Inoculate 2 75 cm2 culture flasks, each with 20 ml of cell suspension of different agronomic levels. Take 0.4 ml per day from the bottles as needed, but mix the cells before sampling. Keep the bottles out of the incubator for as short a time as possible and do not change the culture solution during the growth curve. Alternatively, mount the flasks on an oscillator (Techne, Integra, BeHco) and sample daily. If an oscillator is used to seal the bottle on one side, the bottle can be sampled without removing it from the greenhouse and without opening the sealing film, by wiping the surface of the film clean and turning the bottle upside down to aspirate the liquid with a syringe.