Stampwell U - 3D cell aggregation

Yes, using a 2% agarose solution enables full transparency and therefore direct in situ imaging of your 3D cell aggregates. For optimal imaging in fluorescence and/or confocal microscopy, we recommend transferring your 3D structures into microwells molded using Stampwell V-shape: this stamp has been specifically optimized for parallelized 3D structure imaging by imprinting V-shaped microwells with a flat bottom. 

We recommend using a 2% standard agarose as this was shown efficient to promote 3D structure formation while allowing for proper culture medium diffusion. Yet, you can use your hydrogel of choice to mold your microwells (i.e. Collagen or PDMS). Depending on your experimental goals, rigidity may be tuned to allow proper 3D structure formation, transparency and desired compound diffusion in the gel.   

Yes, once your 3D structures have reached your desired size, they can be collected from the microwells for downstream analysis. Depending on your experimental goals, all aggregates can be flushed out for bulk analysis or individually picked up from the wells. 

Yes, all 3D structures formed are growing separated into individual wells but are sharing the same culture medium. In addition to increasing the number of 3D structures created at once and speeding up the process, this setting prevents them from sticking together while preserving a good viability by maintaining communication between samples. 

So far, Stampwell has been successfully used to make multiple spheroids with HEK, HeLa, U-87 & U2-OS cell lines. We would greatly appreciate your valuable inputs by helping us complementing this list with any of your positive research experiences. Drop us a message through our Contact page!

 Yes, cells can be seeded in Matrigel or other ECM gels in the microwells. You simply need to pre-cool your molded micro-wells at 4°C before seeding, and keep the dish at 4°C while cells are sedimenting at the bottom of the wells before the Matrigel gelation.  

Microwells have a 680 µm diameter and a 517 µm depth.  

The culture medium can be easily replenished in the plate without disturbing the growing 3D aggregates, therefore making it possible to culture them for several days. It is advised not to keep 3D aggregates after they have reached a ~500 µm diameter as bigger structures can be dislodged from the wells more easily. The corresponding time will be very much dependent on the cell type, growth properties and seeding density, but spheroids of desired size are typically obtained after 3 to 4 days. 

Yes, cells can be fixed directly in situ. By pouring liquid agarose on the top, it is then possible to recover fixed 3D structures to perform immunostaining or histological cuts. 

Stampwell has a total external diameter of 26 mm and the microwells are contained in a 20 mm diameter circle. Therefore, it is possible to use the Stampwell in the following plate formats:

• 35 mm circular petri dishes with an aperture of at least 22 mm
• any larger vessel (i.e. 6-well plates)

We provide the stamp to mold microwells. You need to have standard liquid agarose, or the hydrogel of your choice, a petri dish and your cell suspension in your culture medium of choice. 

Molding microwells with Stampwell should take less than an hour. 

Yes, Stampwell U-shape can be reused thousands of times as long as it is properly cleaned and sterilized between experiments (compatible with autoclave sterilization).