A Technique to Isolate Satellite Cells From Rat Head Muscle Tissue

Begin with a rat head muscle and chop it into fragments.

Add a cocktail of proteolytic enzymes and incubate with gentle shaking.

The enzymes digest the tissue's extracellular matrix, loosening the muscle fibers.

Add media containing serum proteins to inactivate the enzymes.

Transfer the tissue to a tube, centrifuge, and discard the supernatant.

Resuspend the tissue in media. Repeat mechanical disruption and centrifugation twice to obtain a homogeneous suspension of satellite cells — stem cells crucial for muscle repair and regeneration.

Collect the satellite cell-containing supernatant.

Pass through a cell strainer to obtain a single-cell suspension. Centrifuge, discard the supernatant, and resuspend the cells in media.

Place drops of the cell suspension onto extracellular matrix protein-coated chamber slide wells.

The coated surface facilitates satellite cell attachment. Add media to the wells.

Growth factors in the media induce satellite cell proliferation and differentiation into myoblasts  — muscle cell precursors, which further proliferate and fuse to form multinucleated myotubes.

The myotubes eventually mature into muscle fibers.