I'm just starting to work with suspension-adapted cells in spinner flasks. and I"m not entirely sure how to go about with their regular maintenance , e.g. passaging, changing media, etc.
does anyone happen to have a pretty straight-forward protocol outlying this??

thanks for the help!!


labrat612

---

It would be useful to know what type and make of spinner flask you are going to use ?

Just some general tips then:

Try and use filtered (0.2uM) tops for your spinner flasks.
Spin at the lowest possible RPM....to prevent cellular damage, but not aggregation.
Optimise the cell density conditions...... my cells are kept between 4,000 and 800,000 cells/ml.
Have plenty of spares for these spinners.....I use Techne stirrers.....spares are glass stirring rods, caps, silicone rubber washers etc etc.

This is usually the easiest way to grow cells..no trypsinisation. Passaging of the cells is just removing some of the cell suspension and adding fresh media, for example:

J774.A1 (murine macrophages): 1ml of cell suspension to 199ml of fresh media.

Hope this is useful.

Kindest regards.

Rhombus

---

Rhombus! that was incredibly helpful.

I'm using 100mL kimble-kontes glass stir flasks and using suspension CHO.

Have you ever tried transfecting suspension cells using the typical reagents: lipofectamine, fugene etc?

---

I have not done any transfection in suspension. I have always been told that transfection rates drop dramatically compared to transfections in adherent cells. Bacullo virus protein expression is the same, dramatic loses when done in suspension.

Regards

Rhombus

---