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A simple, sensitive, selective, precise and stability indicating high-performance thin-layer chromatographic method for determination of gatifloxacin both as a bulk drug and from polymeric nanoparticles was developed and validated as per the International Conference on Harmonization (ICH) guidelines. The method employed thin-layer chromatography (TLC) aluminium plates precoated with silica gel 60F-254 as the stationary phase and the mobile phase consisted of n-propanol–methanol-concentrated ammonia solution (25%) (5:1:0.9, v/v/v). This solvent system was found to give compact spots for gatifloxacin (R_f value of 0.60±0.02). Densitometric analysis of gatifloxacin was carried out in the absorbance mode at 292nm. The linear regression analysis data for the calibration plots showed good linear relationship with r=0.9953 with respect to peak area in the concentration range of 400–1200ngspot⁻¹. The mean …