This chapter reviews the use of the Thin-layer chromatography (TLC) and High performance TLC (HPTLC) in the study of insects. Most of the TLC methods have relied on silica gel for the stationary phase in the analysis of carbohydrates. Most TLC methods for the separation of polyols use silica gel plates. A number of other researchers have used preparative TLC for the fractionation of lipid extracts from insects. TLC has been used in a number of studies for the isolation of ecdysteroids from hemolymph and tissue extracts. TLC has also provided a mechanism to examine the products of ecdysone metabolism. The separation of ecdysteroids by TLC can be accomplished on normal silica gel TLC plates, HPTLC plates, reversed phase TLC and HPTLC plates, and on paraffin–coated silica gel plates. Entomological interest in the terpenoids is primarily in the juvenile hormones, which play a key role in insect development in both the immature and adult stages.