Flavonoids represent a wide group of plant secondary metabolites of intensive interests in recent years because of their many health-related benefits, such as anti-cancer and anti-inflammatory effects. The analysis of these compounds is, however, a complicated one because they are very low in concentration and chemically diverse and have multiple plant metabolites. This chapter gives an overview of the main chromatographic and spectroscopic techniques that are employed in the study of flavonoids, the way they work, their application in research, and future perspectives. The chapter adds the significance of analysis methods in the study of flavonoids, their structure and classification, and challenges associated with their analysis. It also describes various chromatographic methods, including TLC and HPTLC, column chromatography, HPLC, UHPLC, and GC-MS, and spectroscopic and spectrometric methods, including UV-Vis, FTIR, NMR, MS, and fluorescence spectroscopy. The methods also have applications and limitations in flavonoid research, such as studying medicinal plants, herbal products, and biological activity. Finally, the chapter concludes with an inference regarding the state of flavonoid analysis and the requirement for future developments, including the introduction of computer-based laboratories, handheld devices, and green analysis. The overall purpose of this comprehensive chapter is to provide a concise understanding of the advantages and disadvantages of every methodology to the researchers in natural product chemistry, plant-based medicine, and analytical chemistry, so that they can adopt the best methods in their research and effectively utilize flavonoids in medicine.