Cyclosporine A (CsA) is a lipophilic cyclic polypeptide composed of 11 amino acids, seven of which are N-methylated . It has been utilized clinically as a potent immunosuppressant to prevent allograft rejection in various organ transplantations and to treat systemic and local autoimmune disorders [1, 2]. Study by E.Demiryay et al concluded that Opthalmic admistration of Cyclsoporine in artificial tears significantly increases goblet cell density, decreases the signs of DTS (dysfunctional tear syndrome) and improves ocular surface health [3]. Although Cyclosporine ophthalmic emulsion has been on the market for a few years now, exact guidelines on how often and how long dry-eye patients should use it are still debatable. So therefore a high-performance liquid chromatography (HPLC) method was developed and validated to be stability-indicating by stress degradation tests.Due to the cyclic structure and inherent intra-molecular hydrogen bonds, cyclosporine is very stable under normal conditions. However, degradation occurs under stress conditions. Separation of multiple components by HPLC analysis of stability samples possess high accuracy and sensitivity for even small quantities of degradation products produced. In this work, an Acetonitrile-based HPLC method with short run time, with better peak shape and resolution from the excipients has been developed and validated according to the guidance regulated in FDA, and USP. The separation was on a Waters Spherisorb� S5 ODS2 4.6 x 250mm 1.d. reversed phase column at 80� using 80:20 Acetonitrile: water as Mobile phase. Detection was by UV at 210nm, maintained a flow rate of 1mL /min with 20�l injection volume.Three stress conditions were applied: Acid, Alkali, and heat for degradation study. It has been found potential degradation products Isocyclosporine, and Dihydroxy cyclosporine were produced as a result of forced degradation with well resolved and separated peaks from the active indicating successful degradation and stability indicating