Fig. 2

The method of classic “ceiling culture” and its offshoots. A system “ceiling culture”: mature adipocytes (MAs) are seeded in a cell culture flask that is completely filled with medium. After 7 days of cell attachment, the flask was inverted and fresh medium was added to barely cover the bottom of the flask to allow cells to continue to grow; B Preincubation and filter: After a 24-h floating culture period, MAs were transferred to a new petri dish equipped with a 70-μm cell filter. Subsequently, the DFAT cells gradually migrated through the filter and adhered to the bottom of the dish, where they grew and expanded; C Early differential plating: After 1–2 d of ceiling culture, mature adipocytes will be floating in the medium but non–lipid containing cells will attach to the bottom surface. Then floating mature adipocytes in the medium are transferred to the new flask leaving the attached contaminating cells behind; D Late differential plating: Mature adipocytes adhere to the top surface after 3–4 d of ceiling culture. Then the medium is removed and the trypsin of the adherent mature adipocytes are digested and centrifuged, to eliminate the contaminating cells culturing along with mature adipocytes by the buoyant nature of adipocytes. The images presented in Fig. 2 were generated and illustrated by the author