Extracellular vesicles (EVs) play an important role in cell-to-cell communication by carrying molecular messages that reflect physiological and pathological conditions of the parent cells. EVs have been identified in all body fluids; and among them, urine stands out as a sample that is easy and inexpensive to obtain and can be collected over time to monitor changes. Various protocols have been established to study urinary extracellular vesicles (UEVs) and they have shown great potential as a biomarker source for clinical applications, not only for urological, but also non-urological diseases. Due to the high variability and low reproducibility of pre-analytical and analytical methods for UEVs, establishing a standardized protocol remains a challenge in the field of diagnosis. Here, we review UEV studies and present the techniques that are most commonly used, those that have been applied as new developments, and those that have the most potential for future applications. The workflow procedures from the sampling step to the qualitative and quantitative analysis steps are summarized along with advantages and disadvantages of the methodologies, in order to give consideration for choosing the most promising and suitable method to analyze human UEVs.