My experiment aims to make spheroids by sacrificial micromolding technique.
I am following the easy protocol:
"...The PDMS stamps were deposited onto a 250 μl drop of 3% liquid agarose within the wells of a 24-well plate. Agarose gelation occurred within minutes of placing the 24-well plate at room temperature..." Karen E. Samy, 2019.
The PDMS stamp has pillars with 120 um in diameter and 80 um in height according to the article.
However, the problem is, that during pull out of the PDMS stamp the bottoms or walls of agarose microwells are somehow stuck on the pillars, and they are pulled out with them. So, instead of having nice microwells, there are cones made out of pulled microwells, and the cells can't get inside and form a spheroid.
I have tried 3% agarose (according to protocol), 5% agarose, different timing, place the stamps with agarose to fridge or freezer for 10 - 20 minutes, used plasma on PDMS stamps to make them more hydrophilic and all these combinations together.
I am running out of any other ideas. Does anybody know, what could help my situation?
Unfortunately, the corresponding author hadn't answered.
The photo of the "microwells" is attached. In this picture, you see two successful microwells and poorly done the rest of them.