Culturing Salispheres from Salivary Gland Tissue: A Method to Develop Salispheres from Human Salivary Gland Derived Epithelial Cells

Salispheres are in vitro, self-aggregating cultures of epithelial cells derived from the salivary gland. To culture salispheres, begin by taking the human salivary gland tissue in a culture plate.

Using scissors, mince the tissue into fine pieces. Next, supplement these pieces with a dissociation solution containing collagenase and dispase enzymes and incubate. These enzymes digest the extracellular matrix present in the tissue.

Further, triturate to homogenize the tissue. This step releases the epithelial cells and RBCs into the solution. Next, use a cell strainer and filter the tissue homogenate into a fresh conical tube to remove undigested tissue pieces and debris. Centrifuge to pellet the cells.

Following centrifugation, discard the supernatant and resuspend the cell pellet in the RBC lysis buffer. This buffer ruptures RBCs present in the cell suspension.

Now, centrifuge to pelletize the epithelial cells. Resuspend the pellet into an appropriate epithelial cell growth media. Additionally, prepare the salisphere culture plate by coating its well with a liquified basement membrane matrix or BMM. Incubate to allow the matrix to solidify.

Finally, add the epithelial cell suspension into the BMM-coated wells. In the presence of growth media, epithelial cells attach to the BMM layer and proliferate into spherical salispheres.