Hello all,
I'm relatively new to research and just started growing my own Ovarian cancer cells from a pleural effusion.
It's been about 2 weeks now since I've had them in a flask and have changed the media about twice a week. At first the growth was extremely slow, I suspect because of the enviromental change?
However, withing the last few days they began to proliferate much quicker than the previously and now I see one particularly interesting mass and I have no idea if it's just a collection of dead cells or if it's something else. It looks nothing like the regular ovarian cancer cells nor the fibroblasts so not sure if it is and I'm just mistaken.
If anyone could tell me what this is I'd greatly appreciate it!
As you've provided a nice big clear image of what the cells look like that's fairly easy to answer. That is just a big clump of cells that are growing on top of each other. It's a bit easier to see if you adjust the contrast a bit like I have. You can see single cells that you're expecting to see then if you look carefully around the edges in particular where I've drawn white rectangles you can see these cells haven't spread out as much and the further in you go the less spread out they become until it just looks like a shapeless mass.
Cells only spread out like what you typically see in 2D culture on flat hard substrates. On very very soft substrates or in 3D culture they adopt a more rounded appearance. You can see that here, the ones on the very outside have a single cell underneath them they are adhering to between them and the plastic culture dish. They can still feel that hard substrate but because there is a soft cell between them they don't spread out as much. The further into the clump you go the more cells there are, they feel a softer substrate beneath them and there's also less room for them to spread out so they have a more blob like structure.
There is nothing you can (or should) do at this point but when they become confluent enough to split and you've got them in suspension in a falcon tube pipette them up and down 20-30 times with a 1ml pipette. If you use slow deliberate strokes it provides enough shear force to break up cell clumps into more of a single cell suspension without damaging them too much. This should help prevent clumping like you've got here.
As you've provided a nice big clear image of what the cells look like that's fairly easy to answer. That is just a big clump of cells that are growing on top of each other. It's a bit easier to see if you adjust the contrast a bit like I have. You can see single cells that you're expecting to see then if you look carefully around the edges in particular where I've drawn white rectangles you can see these cells haven't spread out as much and the further in you go the less spread out they become until it just looks like a shapeless mass.
Cells only spread out like what you typically see in 2D culture on flat hard substrates. On very very soft substrates or in 3D culture they adopt a more rounded appearance. You can see that here, the ones on the very outside have a single cell underneath them they are adhering to between them and the plastic culture dish. They can still feel that hard substrate but because there is a soft cell between them they don't spread out as much. The further into the clump you go the more cells there are, they feel a softer substrate beneath them and there's also less room for them to spread out so they have a more blob like structure.
There is nothing you can (or should) do at this point but when they become confluent enough to split and you've got them in suspension in a falcon tube pipette them up and down 20-30 times with a 1ml pipette. If you use slow deliberate strokes it provides enough shear force to break up cell clumps into more of a single cell suspension without damaging them too much. This should help prevent clumping like you've got here.
this strongly resembles a patch of immune cells that will grow temporarily: monocytes, B cells and T cells. These cells have been activated by the tumor cell at the beginning of growth, but they will very quickly disappear over the splits of the cell layer.
Howard Vindin
Wow that's much more detailed of an answer than I was expecting. Thank you so much! It helps a lot and I'm sure it will for future reference as well.
Jean-Jacques Pin
Thank you as well for your answer! If they are monocytes then I hope they die off eventually.
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