This study investigates the dynamics and size of lipid rafts in the plasma membrane of mammalian cells using single particle tracking and laser trapping. The researchers measured the local diffusion of raft-associated proteins by confining a microsphere bound to a raft protein in a small area (≤100 nm diameter) using a laser trap. They found that the diffusion of raft-associated proteins is independent of the type of membrane anchor and is significantly reduced compared to non-raft proteins. Cholesterol depletion accelerated the diffusion of raft-associated proteins, bringing their diffusion rates closer to those of non-raft proteins. Raft-associated GPI-anchored proteins remained bound to the raft for up to 10 minutes, indicating that rafts are cholesterol-stabilized complexes of approximately 26 ± 13 nm in size that diffuse as a single entity for minutes. The study also compared the diffusion of proteins with different membrane anchors in intact rafts to those in rafts disintegrated by cholesterol depletion and to non-raft proteins. The viscous drag of raft-associated proteins was found to be independent of the type of membrane anchor and significantly larger than that of non-raft proteins. After cholesterol depletion, the viscous drag of raft-associated proteins decreased to the level of non-raft proteins, while the diffusion of non-raft proteins remained unchanged. The mean radius of the raft assemblies was found to be 26 ± 13 nm. The results support the model that lipid rafts are cholesterol-stabilized complexes that diffuse as a single entity in the plasma membrane. The viscous drag of raft-associated proteins was found to be significantly larger than that of non-raft proteins, indicating that raft-associated proteins are anchored to a membrane patch with a larger diameter than the transmembrane domain of the LDL receptor. The study also showed that raft proteins remain associated with the raft for at least 1 minute, and that rafts diffuse as one entity, with the diffusion coefficient dominated by the motion of the entire raft. The findings suggest that lipid rafts are stable structures that play a role in cell signaling and protein sorting.This study investigates the dynamics and size of lipid rafts in the plasma membrane of mammalian cells using single particle tracking and laser trapping. The researchers measured the local diffusion of raft-associated proteins by confining a microsphere bound to a raft protein in a small area (≤100 nm diameter) using a laser trap. They found that the diffusion of raft-associated proteins is independent of the type of membrane anchor and is significantly reduced compared to non-raft proteins. Cholesterol depletion accelerated the diffusion of raft-associated proteins, bringing their diffusion rates closer to those of non-raft proteins. Raft-associated GPI-anchored proteins remained bound to the raft for up to 10 minutes, indicating that rafts are cholesterol-stabilized complexes of approximately 26 ± 13 nm in size that diffuse as a single entity for minutes. The study also compared the diffusion of proteins with different membrane anchors in intact rafts to those in rafts disintegrated by cholesterol depletion and to non-raft proteins. The viscous drag of raft-associated proteins was found to be independent of the type of membrane anchor and significantly larger than that of non-raft proteins. After cholesterol depletion, the viscous drag of raft-associated proteins decreased to the level of non-raft proteins, while the diffusion of non-raft proteins remained unchanged. The mean radius of the raft assemblies was found to be 26 ± 13 nm. The results support the model that lipid rafts are cholesterol-stabilized complexes that diffuse as a single entity in the plasma membrane. The viscous drag of raft-associated proteins was found to be significantly larger than that of non-raft proteins, indicating that raft-associated proteins are anchored to a membrane patch with a larger diameter than the transmembrane domain of the LDL receptor. The study also showed that raft proteins remain associated with the raft for at least 1 minute, and that rafts diffuse as one entity, with the diffusion coefficient dominated by the motion of the entire raft. The findings suggest that lipid rafts are stable structures that play a role in cell signaling and protein sorting.