Preparation of 3D Differentiation Working Solution

Cell Pretreatment

Pelletization
-If you need to precisely control the number of cells in each iPSC sphere, you can use the microtiter plates provided by Soma Biotech for seeding;
-If the goal is simply to form iPSC spheroids, seed the cell suspension in a 1:1 ratio into non-adherent six-well plates and incubate overnight at 90 rpm on a horizontal shaker in a 37°C incubator.

The next day, we examined the agglomeration results under a microscope and proceeded with the subsequent experiments.

Demonstration of Pellet Formation

Digest a specific amount of iPSCs into single cells and seed them into a honeycomb plate to form spheroids:

A: D0, schematic diagram of cells inoculated individually into a honeycomb plate;

B: Cells that have not been pretreated with 3D differentiation additives form poor spheroids in iPSC maintenance medium;

C: After culturing for 4 days with the 3D differentiation supplement added to the iPSC maintenance medium, the cells were spherocultured using the iPSC maintenance medium; the spheroids formed well, with clear edges and a size within the acceptable range;

D: After culturing for 4 days in iPSC maintenance medium supplemented with 3D differentiation additives, forming spheroids using iPSC maintenance medium plus 3D differentiation additives yields better results, with clearer edges and larger spheroids.