Abstract: A closed-system (i.e., hermetically sealed) methods, and kits for isolating Mesenchymal Stromal Cells (MSCs) from lipoaspirate, that utilizes specific hermetically-sealed, disposable assemblies and containers that are aseptically interconnected, are disclosed. Contemplated methods require lipoaspirate as a starting material, and provide for obtaining isolated, purified MSCs at the end of the process. Kits are contemplated to contain disposable assemblies, composed by sterile components such as modified syringes, tubing, centrifuge tubes, filtering units, that can be aseptically connected while maintaining sterility thereby keeping the system closed (i.e., hermetically sealed) with respect to the external environment. As a result, contamination during the isolation process can be avoided. The MSCs that are obtained at the end of the processing are thus ready to be further manipulated in subsequent operations (in example, expansion) also for therapeutic purposes.