The study investigates the diffusion of phospholipids in the cell membrane, which is significantly slower than in artificial bilayers. Using single-molecule tracking at a high temporal resolution, the authors found that phospholipids are confined within 230-nm compartments for 11 ms before hopping to adjacent compartments. These compartments are further organized into larger 750-nm compartments. The diffusion rate within the 230-nm compartments is comparable to that in large unilamellar vesicles, indicating that the slowdown is due to compartmentalization rather than reduced diffusion rates. The compartmentalization is dependent on the actin-based membrane skeleton, not on the extracellular matrix, extracellular domains of membrane proteins, or cholesterol-enriched rafts. The authors propose that transmembrane proteins anchored to the actin-based membrane skeleton act as "pickets," confining phospholipids through steric hindrance and circumferential slowing. This model explains the double compartmentalization observed in the cell membrane and suggests that the actin-based membrane skeleton plays a crucial role in regulating lipid diffusion.The study investigates the diffusion of phospholipids in the cell membrane, which is significantly slower than in artificial bilayers. Using single-molecule tracking at a high temporal resolution, the authors found that phospholipids are confined within 230-nm compartments for 11 ms before hopping to adjacent compartments. These compartments are further organized into larger 750-nm compartments. The diffusion rate within the 230-nm compartments is comparable to that in large unilamellar vesicles, indicating that the slowdown is due to compartmentalization rather than reduced diffusion rates. The compartmentalization is dependent on the actin-based membrane skeleton, not on the extracellular matrix, extracellular domains of membrane proteins, or cholesterol-enriched rafts. The authors propose that transmembrane proteins anchored to the actin-based membrane skeleton act as "pickets," confining phospholipids through steric hindrance and circumferential slowing. This model explains the double compartmentalization observed in the cell membrane and suggests that the actin-based membrane skeleton plays a crucial role in regulating lipid diffusion.