Self-assembled mesoporous structures with well-ordered nano- scale channels could be used in applications such as molecular separation, nano-optics, molecular electronics, nanomedicine and catalysis1–7. However, the domain sizes that can be created in such systems are limited by our lack of a detailed understanding of the relevant growth processes8–12. Here we report the real-time observation of domain growth in the self-assembly of silica nanochannels using fluorescence polar- ization imaging and atomic force microscopy. We show that transient lamellar structures precede the formation of hexagonal layers, and that the layer growth follows two distinct pathways. In addition, the domains are grown on a mesoporous film substrate, which acts as a sieve and allows control of the delivery of the reactive species. We use these insights and capabilities to grow layers of well-ordered silica nanochannels with domain sizes of up to ~0.3 mm.