The area of application of electrospray ionization mass spectrometry (ESI-MS), usually coupled with liquid chromatography, seems to be much larger than those of any other ionization method. It gives rise to the demand for new teaching methods that would effectively help students to master the principles of working with and making full use of the capabilities of ESI-MS. Herein, a simple demonstration of electrospray ionization mass spectrometry of nucleosides has been proposed in order to show students the basics and capabilities of this ionization method. Besides the simple molecular mass determination followed by analyzing the collision induced dissociation (CID) process, such issues as detection of supramolecular self-assemblies, fragmentation of glycosides, proton/metal cation affinities, and isotope labeling have been discussed on the basis of the mass spectra obtained.