Lignin is the second most abundant plant biopolymer after cellulose on the planet. This phenolic polymer is a major constituent of the cell wall in certain specialised plant tissues where it plays a vital role in many aspects of plant growth. Both the amount and the chemical composition of lignin in cell walls have important effects on the characteristics of plant biomass thereby impacting strongly on the quality of a wide range of economically important products derived from plants. A better understanding of how the lignin polymer contributes to the development of the plant interacts with the environment during the carbon cycle and influences the characteristics of lignocellulose biomass depends upon the availability of appropriate analytical tools permitting a complete characterisation of lignin composition. While several chemical and physical techniques are able to provide both qualitative and quantitative analyses of the chemical composition of extracted cell wall material and/or lignin, they are generally unable to provide detailed spatial information at the tissue/cell-wall level. Recently, the development and application of vibrational spectroscopy and bioorthogonal chemical approaches to lignin imaging are allowing us to obtain a better understanding of the heterogeneity and dynamics of lignin structure in situ. In this article, we discuss lignification and the different techniques that are available to the plant biologist interested in characterising lignin structure with a focus on new imaging techniques.