SK-N-SH detaching - (Aug/13/2008 )

Hi folks,

I got problems with washing SK-N-SH cells on a six well plate with PBS and HBSS. Simply, they detach while washing twice ;-).
Does anyone have any cues to overcome this problem (centrifugation? special plastic ware? I am using the Nunc plates with Nunclon surface)?
Thank you very much!

Regards,

Astat

Dear Astat

I don't know your cell line, but I have experienced similar problems with other cell lines. My conclusions was:

1) My cells detached if the mechanical stress of washing was to big. I solved this by pipetting the medium/PBS very gently on the cell layer or when grown in flasks, simply pipetting medium/PBS onto the side of the flask instead.

2) My cells detached when they became overly confluent. I was following cell cultures over time to follow cell growth and differentiation, and my cells kept detaching half way through my experiments. I solved this problem by seeding fewer cells at the onset of the experiment. I did a small pilot seeding cells at 1000, 3000, 6000, 12000 and 20000 cells/cm2. For my cells a seeding density of 6000 cells/cm2 was the best.

I hope this gives you an idea, how you can solve your problem. There are other options, for example coating the wells with matrigel or similar -but this tend to make routine work very complex.

Best regards

Kirsten