I would propose to use a 2-chamber system (e.g. see CytoSelect™ 24-Well Cell Co-Culture System (Cell Biolabs; Cat# CBA-160, for protocol see: https://www.cellbiolabs.com/sites/default/files/CBA-160-cell-co-culture-system.pdf).

For additional systems, e.g. see the following review article:

Goers L, Freemont P, Polizzi KM. J R Soc Interface. 2014 Jul 6;11(96). pii: 20140065. doi: 10.1098/rsif.2014.0065.

What people usually do is they first grow the cancer cells (feeder cells), and then add the other cells on top of them so they can get feeded by these cancer cells

But since in your case both of them are cancers cells, you can transfect Jurkat first then trypsinize both of cells in same day and mix them together

Sorry I mean trypsinze the adherent cells only

Jurkat already floating cells so no need for trypsin

Sure, that`s a standard procedure (as mentioned in every hand book of any adequte assay). However, in any case you should NOT do this in the same well. You should use a system allowing discriminating between both cell types (as mentioned above). I would give the same advice to Abdullah!

Culturing the two different types of cancer cells together shouldn't be a problem. The real question is, 'How should you do it so that you can observe what you need to observe?"

If your Jurkat are transfected, they probably need a selection agent/antibiotic of some sort, do they not?

Well, Are the adherent cells also transfected so that they have the gene for resistance to the selection reagent?

Or are you Jurkat cells 'transformed'? In this case, no selection agent is required, however, they will lose the dna they are transformed with in a matter of days. This narrows your window of opportunity and should be considered when working up your protocol.

Also,

• Do you need to adherent cells to be near confluent?
• Do you need to put them through some sort of Serum Starvation for a short time to synchronize them?
• Do you plan to observe receptor ligand interactions between the cells? If so, will you be using a confocal microscope? If so, will the cells be plated on glass cover-slips which have been coated with protein to allow the adhesion cells to adhere?

There's a lot to consider...

Can you give us any more information to go on?

I think what you're doing sounds interesting. I'd like to hear more about it.

Thanks all for your comments and suggestions!

Tim Tolentino: I used Jurkat as a model T cells to study cancer-triggered activation of immune cells. That being said, my Jurkat cells were transfected with a receptor construct that can recognize specific cancer cell antigen. When co-culture Jurkat and cancer cells, I would access T cell's activation through functional output tests (e.g. secretion of IL-2/IFN-gamma cytokine, expression of activation marker CD69, etc.).

So I was wondering how to properly co-culture them and how to force the contact of the cells in co-culture while cancer cells are adherent and Jurkat are suspension cells.

That sounds like a great experiment.

Have you considered using a temperature controlled flow chamber or something similar?

There are probably a lot of different chambers out there now but basically the setup is a closed chamber in which the bottom is a cover-slip onto which adhesion cells have been cultured (the cover slips are treated with poly-D lysine and ECM to allow adhesion). Since the chamber is a flow chamber, solutions can be injected and the excess flows out the exit. As such, suspension cells (like Jurkat cells) can be injected into the chamber. Once the suspension cells are in the chamber, the chamber is sealed (turn the stopcock) and the suspension cells can then interact with the adhesion cells...

Now, because the adhesion cells are cultured onto a glass cover-slip, you can have this chamber on an inverted microscope and you can observe what is happening inside the chamber. Also, because, the chamber is temperature controlled, the cells are kept at physiologic temperature and able to go through their normal processes.

So, you have a way to:

1: force contact between the Jurkat cells and the adhesion cells

2: observe the interaction

3: fluorescently label the different markers you might be interested in...

You could capture live 3D images over time of the cells interacting.

If you think this might work for you, I'd be happy to go through the details with you. Just send me a message.

Good luck!