High-performance liquid chromatographic (HPLC) analysis of antiretroviral (ARV) drugs in biological matrices are reviewed for therapeutic drug monitoring purposes. HPLC is the most often used analysis method of these drugs. Liquid-liquid extraction, solid phase extraction and protein precipitation were used for the prepurification of the biological samples. ARV drugs were detected by various detection methods, including ultraviolet, fluorescence, mass spectrometry ( MS) and MS/MS. This review shows that HPLC methods allow quantitative determination of antiretroviral drugs, individually or simultaneously in biological matrices for therapeutic drug monitoring purposes.