A sensitive, reliable, and rapid densitometric high-performance thin-layer chromatography (HPTLC) method has been developed for the quantification of hydroxysafflor yellow A (HSYA) in safflower. Chromatographic analysis was performed using the methanol extract of safflower. Polyamide TLC plates were used with the solvent system containing 3.6% hydrochloric acid, methanol, and ethyl acetate (7:3:1, v/v). Densitometric analysis of HSYA was performed at the absorbance mode of 399 nm. This system was found to give compact and dense spots for HSYA (RF value of 0.60 ± 0.03). The method showed a good linear relationship in the range of 61.0– 79.3 ng with r = 0.9991. The limit of detection, limit of quantification, and average recovery were 59 ng, 169 ng, and >95%, respectively. The proposed validated HPTLC method is an easy-to-use, accurate, and convenient method that can be successfully used for the standardization and quality analysis of herbal materials with different species of safflower.