Molecular self-assembly is a fundamental process in nature that can be used to develop novel functional materials for medical and engineering applications. However, their complex mechanisms make the short-lived stages of self-assembly processes extremely hard to reveal. In this article, we track the self-assembly process of a benchmark system, double-walled molecular nanotubes, whose structure is similar to that found in biological and synthetic systems. We selectively dissolved the outer wall of the double-walled system and used the inner wall as a template for the self-reassembly of the outer wall. The reassembly kinetics were followed in real time using a combination of microfluidics, spectroscopy, cryogenic transmission electron microscopy, molecular dynamics simulations, and exciton modeling. We found that the outer wall self-assembles through a transient disordered patchwork structure: first, several patches of different orientations are formed, and only on a longer time scale will the patches interact with each other and assume their final preferred global orientation. The understanding of patch formation and patch reorientation marks a crucial step toward steering self-assembly processes and subsequent material engineering.

Self-assembling amphiphilic cyanine dyes, such as C8S3, are promising candidates for energy storage and optoelectronic applications due to their efficient energy transport properties. C8S3 is known to self-assemble in water into double-walled J-aggregates. Thus far, the molecular self-assembly steps remain shrouded in mystery. Here, we employ a multiscale approach to unravel the first self-assembly step: dimerization. Our multiscale approach combines molecular dynamics simulations with quantum chemistry calculations to obtain a Frenkel exciton Hamiltonian, which we then use in spectral calculations to determine the absorption and two-dimensional electronic spectra of C8S3 monomer and dimer systems. We model these systems solvated in both water and methanol, validating our model with experiments in methanol solution. Our theoretical results predict a measurable anisotropy decay upon dimerization, which is experimentally confirmed. Our approach provides a tool for the experimental probing of dimerization. Moreover, molecular dynamics simulations reveal that the dimer conformation is characterized by the interaction between the hydrophobic aliphatic tails rather than the π–π stacking previously reported for other cyanine dyes. Our results pave the way for future research into the mechanism of molecular self-assembly in similar light-harvesting complexes, offering valuable insights for understanding and optimizing self-assembly processes for various (nano)technological applications.

Microfluidic mixing captures self-assembly from disordered molecular assemblies to well-structured double-walled nanotubes, with characteristic optical signatures resolved by two-dimensional electronic spectroscopy.

Self-assembly, the spontaneous organization of molecular components into ordered structures without external intervention, offers a powerful route to complex nanomaterials, yet the molecular-level pathways, particularly under non-equilibrium conditions, remain poorly understood. We track in real time the formation of double-walled nanotubes (DWNTs) from the amphiphilic cyanine dye C8S3, a synthetic analogue of light-harvesting chlorosomes in green sulfur bacteria. Combining microfluidic control with two-dimensional electronic spectroscopy, cryogenic transmission electron microscopy, and polarization-resolved single-object microscopy, we identify transient intermediates at both structural and excitonic levels. The results show that the outer nanotube structures first, while the inner nanotube follows with a delay, ultimately giving rise to electronically coupled coaxial architectures. While principal excitonic and morphological features emerge within minutes of self-assembly, axial elongation and orientational refinement proceed over tens of hours in a nucleation–elongation fashion. Notably, suppressing local concentration gradients through more efficient mixing abolishes DWNT formation, establishing spatial heterogeneity as a key design parameter for hierarchical self-assembly. These findings provide a framework for controlling biomimetic light-harvesting nanomaterials, with implications for directing structural outcomes in advanced materials engineering.