Linezolid is one of the first commercially available and most widely used oxazolidinone antibiotics. This study describes the development of a microbiological assay, applying the turbidimetric method for the determination of linezolid, as well as the evaluation of the ability of the method in determining the stability of linezolid against acidic, basic, neutral, thermal and photolytic degradants using Staphylococcus aureus ATCC 6538P as the test micro-organism. The results obtained with the microbial method were compared with the results obtained from the spectrophotometric method, developed and validated using ferric citrate and 1, 10-phenanthroline reagents to give an orange coloured chromogen, measured at 510 nm. The MIC value for standard linezolid was found to be 2.7 μg/ml. It was observed that linezolid is unstable in acid and alkali. The potency of linezolid is critically reduced under these stress conditions, whereas under neutral, thermal and photolytic stress conditions the potency of linezolid was reduced to a smaller extent.