Liquid chromatography-tandem mass spectrometry (LCMS/MS) is one of the most advanced techniques in the development of Methods for estimation of drugs in biological fluids. The current work includes basic techniques in Liquid Chromatography, Mass Spectrometry, Sample Preparation in bioanalyses, development of bioanalytical method for the quantitative estimation of Montelukast in human plasma and validating it according to FDA guidelines. The method finds its application in biequivalence studies of Montelukast, a cysteinyl leukotriene receptor antagonist used in the treatment of asthma and other allergic conditions.