Membrane microdomains enriched in cholesterol, sphingolipids (rafts), and specific proteins are involved in important physiological functions. However their structure, size and stability are still controversial. Given that detergent-resistant membranes (DRMs) are in the liquid-ordered state and are rich in raft-like components, they might correspond to rafts at least to some extent. Here we monitor the lateral order of biological membranes by characterizing DRMs from erythrocytes obtained with Brij-98, Brij-58, and TX-100 at 4 °C and 37 °C. All DRMs were enriched in cholesterol and contained the raft markers flotillin-2 and stomatin. However, sphingomyelin (SM) was only found to be enriched in TX-100-DRMs – a detergent that preferentially solubilizes the membrane inner leaflet – while Band 3 was present solely in Brij-DRMs. Electron paramagnetic resonance spectra showed that the acyl chain packing of Brij-DRMs was lower than TX-100-DRMs, providing evidence of their diverse lipid composition. Fatty acid analysis revealed that the SM fraction of the DRMs was enriched in lignoceric acid, which should specifically contribute to the resistance of SM to detergents. These results indicate that lipids from the outer leaflet, particularly SM, are essential for the formation of the liquid-ordered phase of DRMs. At last, the differential solubilization process induced by Brij-98 and TX-100 was monitored using giant unilamellar vesicles. This study suggests that Brij and TX-100-DRMs reflect different degrees of lateral order of the membrane microdomains. Additionally, Brij DRMs are composed by both inner and outer leaflet components, making them more physiologically relevant than TX-100-DRMs to the studies of membrane rafts.

富含胆固醇，鞘脂（筏）和特定蛋白质的膜微区参与重要的生理功能。然而，它们的结构，尺寸和稳定性仍存在争议。假设耐洗涤剂的膜（DRM）处于液体有序状态且富含筏状成分，则它们可能至少在一定程度上与筏相对应。在这里，我们通过表征在4°C和37°C下用Brij-98，Brij-58和TX-100获得的红细胞的DRM来监测生物膜的横向顺序。所有DRM均富含胆固醇，并包含筏标记flotillin-2和stomatin。但是，鞘磷脂（SM）仅在TX-100-DRM（一种优先溶解膜内小叶的去污剂）中富集，而Band 3仅存在于Brij-DRM中。电子顺磁共振光谱显示，Brij-DRMs的酰基链堆积低于TX-100-DRMs，为它们的不同脂质组成提供了证据。脂肪酸分析表明，DRM中的SM部分富含木素酸，这应特别有助于SM对洗涤剂的抵抗力。这些结果表明，来自外部小叶的脂质，特别是SM，对于DRM的液相有序相的形成至关重要。最后，使用巨型单层囊泡监测Brij-98和TX-100诱导的差异增溶过程。这项研究表明，Brij和TX-100-DRM反映了不同程度的膜微区横向顺序。此外，Brij DRM由内部和外部小叶组件组成，