Cystic fibrosis is a life-threatening genetic disease that causes damage to the lungs. Ivacaftor, the first drug to target the underlying defect of the disease caused by specific mutations, improves outcomes and reduces hospitalizations. In this study, quantitative determination of ivacaftor was performed by liquid chromatography, while high-resolution mass spectrometric analyses were performed for qualitative determination. The validation studies of the developed methods were performed according to International Conference on Harmonisation Q2(R1) guideline. Ivacaftor was separated from its degradation product by using Phenomenex Kinetex C-18 (150 x 3 mm, 2.6 mu m) column. The isocratic mobile phase for binary pump configuration was 0.1% (v/v) formic acid in water and 0.1% (v/v) formic acid in acetonitrile (27:63) (v/v), pH = 2.5; the flow rate of 0.25 mL/min was used in all methods. In the degradation studies, five degradation products were identified using high-performance liquid chromatography ion trap time-of-flight mass spectrometric analyses: three of them have never been reported up to date; whereas the other two were existing in the literature and they were having Chemical Abstracts Services registry numbers since they were synthesized before for various other purposes. Also, analysis of an in-lab prepared chemical equivalent of Kalydeco (R) and interlaboratory comparison were performed.