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Flavonoids are a large group of plant secondary metabolites that are present in most plants, and are vital for plant growth, development and protection. Among the many functions of these compounds is their contribution to stress amelioration. The accurate identification and quantification of total or individual flavonoids in plants exposed to stressful conditions (e.g. ultraviolet radiation) is challenging due to their structural diversity. The present review provides the up to date knowledge and highlights trends in plant flavonoid analysis. The review covers all steps from the field to the laboratory, focussing on UV-B effects on flavonoids, and identifying critical issues concerning sample collection, pre-treatment, and extraction techniques and quantitative or qualitative analysis. A well-planned sampling and sample prehandling strategy is vital when capturing organ, tissue and developmentalstage dependent changes in flavonoids, as well as the dynamic changes due to time of UV-exposure and diurnal or seasonal parameters. A range of advanced extraction and purification techniques can facilitate the quantitative recovery transfer of flavonoids to solvents. The advantages and disadvantages of analytical methods, including chromogenic assays, liquid and thin-layer chromatography, mass spectrometry, NMR detection, and non-destructive in situ fluorescent analysis need to be consciously evaluated in the context of the specific biological question posed. Thus, no one method can be applied to every single study of flavonoid. The message of this review is that researchers will need to carefully consider the biological process that they intend to study, and select an analytical method that optimally matches their specific objectives.