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Septin family proteins oligomerize through GTP-binding domains into core heteromers, which in turn polymerize at the cleavage furrow of dividing fungal and animal cells. Septin assemblies during the interphase of animal cells remain poorly defined and are the topic of this report. Here we developed protocols for visualization of authentic higher-order assemblies using tagged septins to effectively replace the endogenous gene-product within septin core heteromers in human cells. Our analysis revealed that septins assemble into microtubule-supported disc-like structures at the plasma membrane. In the absence of cell substrate-adhesion, this is the predominant higher-order arrangement in interphase cells and each one of the 7 to 8 septin family members expressed by the two analyzed cell types appears equally represented. However, studies of myeloid and lymphoid cell model systems revealed cell type specific alterations of higher-order septin arrangements in response to substrate-adhesion. Live-cell observations suggested that all higher-order septin assemblies are mutually exclusive with plasma membrane regions undergoing remodeling. The combined data point to a mechanism by which densely arranged cortical microtubules, which are typical for non-adhered spherical cells, support plasma membrane-bound disc-like septin assemblies.