Genome duplication is essential for cell proliferation. Errors in the mechanisms that control DNA replication can cause genomic instability and lead to the development of genetic diseases and cancer. In vitro reconstitution of DNA replication with purified yeast proteins has helped uncover important mechanisms of DNA replication. How changes in protein structure regulate many key functions during initiation of DNA replication and fork progression remain unknown or speculative. To date, structural studies have focused on imaging artificially isolated replication complexes using simplified DNA substrates to understand DNA unwinding and replisome architecture. To truly understand the mechanisms that control DNA replication, future structural studies must not only visualise isolated complexes, but also reconstituted reactions. Nanobodies targeting short linear peptide-epitopes have become increasingly relevant in structural biology, notably for handling fragile samples. Here, we leverage the ALFA nanobody technology to gain structural insights into origin-dependent eukaryotic CMG assembly, using in vitro reconstituted cellular reactions and electron-microscopy. Our methodology offers an efficient protocol for purifying replication intermediates from low particle concentrations under both native and cross-linked conditions. This surpasses the large amounts of protein of conventional methods like GraFix and overcomes high background signals when imaging entire reactions in solution. Using the CMG formation reaction as proof of concept, we've improved the purification of DNA-bound and protein-protein complexes by up to a factor of 5-fold, while effectively eliminating contaminating excess background. The incorporation of nanobody technologies offers a robust simple tool for researchers to isolate both native and cross-linked stabilised samples from complex biochemical reactions for high-resolution structural studies.