Not sure what you mean by incomplete medium. If it is hypotonic it could be causing the cells to round up by rapid cell swelling.  Alternatively, if there is not sufficient Ca++ and Mg++ the integrins could be inactivated and result in a loss of adhesion.  

Which medium do you use? Neuroblastoma cells growt better in RPMI.

By imcomplete medium you mean without FBS? Or without growth factors?

The medium was DMEM without FBS.

From my experience, this cells need to growth in FBS. This could be the reason why the change its morphology.

Dear Carla, initially the cells are grown in the  presence of FBS only. It's only observed after the medium change to -FBS type for carrying our further experiments.

I've starved SHSY5Y in the past using Serum-free media and did not experience this (even after 18h+). Have you tried preparing the incomplete media again? I'd imagine that this may be due to the incomplete media being hypotonic...

Do the cells exhibit normal morphology if you return them to 'full' media after the incomplete medium?  

Dear Davor,

I have prepared fresh incomplete media and used it, but the problem still persists. When i return them to complete media again after incomplete media, they do not exhibit normal morphology.

Dear Ayushi,

I assume that you are omitting the serum so that the medium does not contain any growth factors.

We should always keep in mind that the cells do not only get growth factors from the serum. There are two things you may want to try.

1) Serum contains albumin. The albumin in the serum protects cells from reactive oxygen species (ROS). Albumin therefore acts as an antioxidant. Albumin serves as a transporter of important biological ligands (probably not important for your situation) and it protects cells against physical damage (hydrodynamic shear forces) when pipetting and centrifuging the cells. So, the rounding up of the cells could be because they are dyeing due to too much ROS or due to physical damage. To solve this, add BSA to the medium (final concentration 1%) and see if this solves your problem.

2) Serum also contains factors (fibronectin, laminin, serum spreading factor) that promotes cell attachment to the culture flask. To address this, you may want to coat the flask with gelatine (0.1-2%).

I hope this solves your problem.

Greetings,

Wynand

Dear Ayushi:

This cell are neuroblastom and cold be differentiated to neuronal phenotype by serum deprivation like as PC12 cells, Cad cells , and Others. Thus is normal that cells change its morphology.  

Please refer to the SHSY5Y Product Sheet information (ATCC® CRL2266™).

" These cells grow as a mixture of floating and adherent cells. The cells grow as clusters of neuroblastic cells

with multiple, short, fine cell processes (neurites). Cells will aggregate, form clumps and float."

Although morphological differentiation can be initiated by serum deprivation, it is a long process that needs some time to kickoff.

The immediate effect on your cells might be due to:

-Roughly handling the cultures which can cause the cells float in the medium. Many of the cells often detach and become suspended in the culture medium.

-Thermal shock to the cells

-Osmotic shock

I would suggest to add BSA 1% to the media, as others suggested. We are used to do this in our lab when FBS is needed to be removed.

Hope it helps!

Hello, as Chiara pointed out already we had also beneficial effects by adding 1% BSA to cultures of epithelial cells in defined ('incomplete') media. However, in our case cells produce and secrete extracellular matrix (ECM) components which are probably missing in your cultures. So you may add fibronectin or vitronectin both of which are ingredients of serum, such as FCS.

I assume you are growing the cells on a substrate coated for cell adhesion. An autoclaved water solution of 0.1% Knox brand unflavored plain gelatin (you can buy it in the grocery store) can be applied to flasks, dishes, plates, wells, coverslips, etc., then removed and air-dried in the hood. This simple treatment provides a more physiological cell substrate and has remarkable effects on cell morphology, viability, etc.

Could you update us on what solved your problem? I'm facing the same issue with all my cell lines (started recently)