A new, simple, sensitive, selective, precise and stability-indicating high-performance thin-layer chromatographic method for analysis of strychnine in Strychnos nux vomica seed extract, and in marketed formulation was developed and validated. The method was developed on TLC aluminum plates precoated with silica gel 6OF254 as the stationary phase. The solvent system consisted of toluene: ethyl acetate: diethyl amine (7: 2: 1, v/v/v). Densitometric analysis of strychnine was carried out in the absorbance mode at 254 nm. This system was found to give compact spots for strychnine (Rf value of 0.42 士 0.01, for six replicates). Strychnine was subjected to acid and alkali and hydrolysis, hydrogen peroxide-induced degradation and photo degradation. The drug undergoes degradation under all stress conditions. Also, the degraded products were well resolved from the pure drug with significantly different Rf values. The method was validated for linearity, precision, robustness, LOD, LOQ, specificity and accuracy. Linearity was found to be in the range of 100-1000 ng/spot with significantly high value of correlation coefficient r2 = 0.9928 士 1.02. The limits of detection and quantification were 12.00 and 36.37 ng/spot, respectively. Statistical analysis proves that the method is repeatable, selective and accurate for the estimation of strychnine in S. nux vomica seed extract, and in market formulation. The developed method effectively resolved the strychnine in S. nux vomica seed extract, and in marketed formulation hence; it can be employed for routine analysis as a stability indicating method.