The formation of infection threads in the symbiotic infection of rhizobia in legumes is a unique, fascinating, and poorly understood process. Infection threads are tubes of cell wall material that transport rhizobia from root hair cells to developing nodules in host roots. They form in a type of reverse tip-growth from an inversion of the root hair cell wall, but the mechanism driving this growth is unknown, and composition of the thread wall remains unclear. High resolution, 3-dimensional imaging of infection threads, and cell wall component specific labelling, would greatly aid in our understanding of the nature and development of these structures. To date, such imaging has not been done, with infection threads being imaged simply by bright field, by membrane labels, and histochemically in thin sections. We have developed new methods of imaging infection threads using novel and traditional cell wall fluorescent labels, and laser confocal scanning microscopy. Of these methods, we have used a new highly intense and stable pseudo-Schiff/rhodamine-123 based label, which has allowed for 3D imaging of infection threads in greater detail than had previously been achieved. Through novel combinations of the above method and calcofluor-white staining, we have succeeded in differentially labelling infection threads, and have visualized separate components of thread cell walls. This differential labelling of infection threads and imaging of whole samples by confocal microscopy has also made possible a more detailed and accurate quantification of thread phenotypes and numbers than had until now been practicable. Our methods have made the imaging and study of infection threads considerably more efficient and informative, and present exciting new opportunities for future research in the area.