This chapter provides an overview of the theory and practice of Thin-Layer Chromatography (TLC) and offers a brief summary of the new advances in this rapidly developing field of chromatographic separation science for researchers and serious students interested in any of the topics covered. TLC was developed more than 30 years ago for the separation and semi-quantitative determination of the individual components of more or less complicated mixtures. TLC plates are prepared from these materials by binding them to a support with the help of various organic or inorganic binders. The position of any solute spot in TLC is characterized by the retention factor Rf. TLC is a suitable chromatographic method for the separation and quantitative determination of solutes that are present in a low quantity in complicated organic and/or inorganic matrices. The linear ascending developing method is the most frequently used in TLC practice.