In support of individualizing antiretroviral treatment by combining therapeutic drug monitoring (TDM) and HIV resistance tests, numerous assays have been reported in the literature in an attempt to optimize treatment outcomes. These assays vary with regard to analysis method of separation and detection such as using high performance liquid chromatography (HPLC) coupled to mass tandem spectrometry (HPLC/MS or HPLC/MSMS) or ultraviolet spectrometry (HPLC/UV). To prepare plasma samples prior to analysis, clinical samples are subject to chemical procedures including solid phase extraction (SPE), liquid-liquid extraction (LLE), and protein precipitation (PP). Information searches were performed using database searching tools such as PubMed, MEDLINE, Web of Science, and EMBASE using the key words protease inhibitors, HIV, assay, quantification, determination, therapeutic drug monitoring and pharmacokinetics. Publications were reviewed and the data categorized by the specific protease inhibitor, the limit of quantification (LOQ), the type of assay, chromatographic conditions, sample preparation, and total assay run time. The analytical methods summarized in the review reflect the timeframe since the PI were introduced, their eventual combination with ritonavir, transitioning toward once or twice daily dosing and more recently their use in simplified regimes for chronic dosing. As antiretrovirals are introduced into developing countries additional considerations related to drug quality and potency may require that laboratories adapt these assays in support of clinical pharmacology research and patient monitoring.