AbstractSelf-assembly of isoporous membranes of polystyrene-b-poly(4-vinylpyridine) (PS-b-P4VP) involves many parameters: the block copolymer composition, the solvent and its interactions with the respective blocks, composition of the casting solution, solvent evaporation, and immersion into a water bath. We characterized the self-assembly of PS-b-P4VP in solution in nearly neutral or P4VP-selective solvent and on the surface of a cast film. We used a combination of room temperature (RT) and cryogenic high-resolution scanning electron microscopy (cryo-HR-SEM) to study copolymer micellization in dried films, in solution, and on the membrane surface during evaporation. The solutions, with and without addition of small water amounts, were investigated by cryogenic transmission electron microscopy (cryo-TEM) and small-angle neutron scattering (SANS). We have shown that the starting point for membrane formation is a microphase-separated copolymer solution with the P4VP within the micelle core. As water is introduced, this structure is preserved as long as the system is far from equilibrium. Closer to equilibrium the PS blocks form the micelles core.