Of course, the media volume and media height, respectively, covering a cell monolayer may have a significant impact. For each culture vessel (flasks, dishes, multiwell plates) the usual volumes are published in tables by the vendors. Normally, culture medium should cover the cells by 2 - 5 mm, corresponding to 0.2 - 0.5 ml/cm2 growth area.

Medium height determines the diffusion distance for oxygen to the cells. I have enclosed a paper, in which we increased the medium volume covering renal epithelial monolayers, which resulted in a significant increase in rate of glycolysis, i.e. less oxidative metabolism due to decreased oxygen supply.

 Hi there,

Good day to you. The volume must be in a way that constantly supply the nutrients to the cells. Having excess media would do the trick, but do bear in mind that their excretory products are indeed toxic to the cells if left unattended.

My suggestion is to check every alternate days and check for media colour change if it's phenol red based. This way you could keep track of your cell growth.

Hope this helps. Thanks.

Regards,

Naresh. 

Ravi,

Are your cells adherent or not? If they are growing in suspension culture, you should keep them in certain limits (such as 2E5 - 1E6 cell/ml). On the other hand, I change the media volume according to the seeding density of the cells for adherent cells, 2 - 5 ml of media for a T25 flask.

Best,

Ozgun

please consider these two issues:

1) reduced medium volume implies minor nutrient/factors availability, and evaporation danger, anyway it's easily detected by pH indicator color changing;

2) excess medium volume it's safe against nutrient depletion, but it can limit gas (CO2 atmosphere) and heat exchange...

you have to observe, evaluate and learn the best condition for your specific culture, to manage medium volumes.

Hi Ravi,

For all types of flaks and multi wells there is an optimum medium volume. If you culture with more medium, gas exchange will be the biggest problem and this can influence the cell behavior. When the cell density is not so high you can change the medium once a week

best,

Cor

Of course, the media volume and media height, respectively, covering a cell monolayer may have a significant impact. For each culture vessel (flasks, dishes, multiwell plates) the usual volumes are published in tables by the vendors. Normally, culture medium should cover the cells by 2 - 5 mm, corresponding to 0.2 - 0.5 ml/cm2 growth area.

Medium height determines the diffusion distance for oxygen to the cells. I have enclosed a paper, in which we increased the medium volume covering renal epithelial monolayers, which resulted in a significant increase in rate of glycolysis, i.e. less oxidative metabolism due to decreased oxygen supply.

Another issue is that a good many cell types secrete autocrine growth factors that they actually need.  So if you add too much medium then they cannot sustain a high enough level of factors and may die.  The same applies to small MW nutrients like thymidine - if there are enough cells then they can condition the medium with these nutrients but if there is too much medium then they can't.   So it is possible to add either too much or too little medium - as someone said, you just have to find out the best level for your cells.  Attached normal(ish) cells typically prefer to be above 3-5x104 /ml and up to about 3-6 x 105/ml (it varies).   A rough guide (regarding "too little")  is to change the medium when the colour starts to change towards orange.  

By the way - medium (singular); media (plural).

It depends on the cell type - but the general rule is that you seed the cells at 1x10^5/ml  to 1x10^6/ml, then you should observe them everyday to monitor the growth of the cells - if they are happy or not, then you can adjust the concentration to optimize the condition. 

Medium volume must be determined through the cell type. Generally you can feed cells with app. 10 ml in 75 cm2 flasks and app. 5ml in 25 ml flasks. It is necessary for diffision of oxgen and other nutrients.

Yes volume of medium affects cell growth;good idea to check previous publications, for specific cell types, but if you are using individual glass bottom dishes for growing neurons for example, you can have the benefit of a large volume (2 - 3 ml) spread out over a greater surface area thereby reducing the thickness that would otherwise compromise gas exchange (as in a multiwell plate). Hope this helps.

The volume of cell culture media is very important to optimize experimental conditions. The amount of media affects the osmolality of cultured cells. Osmolality increases during the cell culture. This procedure is regulated by the solubility of oxygen and CO2 (as a buffering system). Threfore, optimized amount of media should be used for selected tissue culture vessels.

Best wishes

In my practical experience I have seen that human umbilical vein endothelial cells (HUVEC) grows best if I put 1x 105 ( one into ten to the power five) cells in a 100mm plate with a 10 ml culture medium (M199). The cell will not grow if I put less cell in the culture medium. Also instead of 10 ml if I put 20-25 ml medium that actually somehow may slightly decrease the growth rate. Definitely increasing the cell culture medium from 10 ml to 25 ml in 100 mm plate does not have any additive beneficial effects.