Preparation of Cancer Spheroid Blocks by Flash Freezing: A Procedure To Preserve Cancer Spheroids in Frozen Matrix Blocks

First, use small forceps to lift the block of collagen gel and spheroids from the chamber slide. Then, place the collagen gel block in a plastic histology mold. Rinse the collagen gel block with 1X PBS briefly and fix it with 4% PFA in PBS for 30 minutes at room temperature. After this, wash the collagen gel block with 1.5 milliliters of 1X PBS on a shaker for 15 minutes.

Apply a thin layer of OCT compound to the bottom of a new plastic histology mold. Then, place the fixed and washed collagen gel block on top of the OCT compound before carefully filling the rest of the mold with OCT compound. Keep the mold at 4 degrees Celsius for 1 hour. After this, flash-freeze the mold on a 100-millimeter Petri dish floating on liquid nitrogen, and store the samples at minus 80 degrees Celsius.