I need to fix the 3D tumor spheroid that has been constructed using the low attachment plate method for confocal imaging. We are having problems with fixation without changing the sphere shape. Does anyone know a better method to overcome this issue?
You can fix the spheroids using 4% PFA or Formaldehyde for 20 to 30 min @ RT and kept stored in PBS at +4 C and they are fine for 1-2 months without changing their shape. Each cells are different and their fixative buffer and fixation time differs from each other.
And for the transportation of the spheroids you can use Matrigel or you can use gelatin (thin layer) coated over the glass plate or cover slides and then culture the spheroids of your interest. This prevents the change in shape of the spheroids also helps in better attachment of the cells.
For transportation, fix them with 4% PFA (20-30 mins @RT) and then paraffin embed. you can store them for various types of histological analysis and immunofluorescence too.
Hi Janith, please let me know how did you solve the problem? I am struggling to image the spheroid as it is moving
@Florian Kai Groeber-Becker Could you please give me more detail on how to embed the fixed spheroids in Low melting agarose
Cheers
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