Magnetic alignment of rhodamine/magnetite dual-labeled microtubules probed with inverted fluorescence microscopy

Molecular machines, as exemplified by the kinesin and microtubule system, are responsible for molecular transport in cells. The monitoring of the cellular machinery has attracted much attention in recent years, requiring sophisticated techniques such as optical tweezers, and dark field hyperspectral and fluorescence microscopies. It also demands suitable procedures for immobilization and labeling with functional agents such as dyes, plasmonic nanoparticles and quantum dots. In this work, microtubules were co-polymerized by incubating a tubulin mix consisting of 7 biotinylated tubulin to 3 rhodamine tubulin. Rhodamine provided the fluorescent tag, while biotin was the anchoring group for receiving streptavidin containing species. To control the microtubule alignment and consequently, the molecular gliding directions, functionalized iron oxide nanoparticles were employed in the presence of an external magnet field. Such iron oxide nanoparticles, (MagNPs) were previously coated with silica and (3-aminopro-pyl)triethoxysilane (APTS) and then modified with streptavidin (SA) for linking to the biotin-functionalized microtubules. In this way, the binding has been successfully performed, and the magnetic alignment probed by Inverted Fluorescence Microscopy. The proposed strategy has proved promising, as tested with one of the most important biological structures of the cellular machinery.