Endoscopic ultrasound-guided fine needle aspiration biopsy (EUS-FNA) is currently performed on a routine basis at many endoscopic centers and it is evident that this procedure has a major impact on the therapeutic management of patients by obtaining a definite tissue diagnosis from lesions outlined by endosonography. The reported yield of EUS-FNA is about 90-95%, with an overall sensitivity and specificity of 90% and 100%, respectively. Moreover, even minute lesions down to a size of 5 mm may be imaged and consequently biopsied. This Review describes the technique of EUS-FNA in detail, based on a literature review and the authors' extensive experience with this method. The endoscopes and needle systems available on the market are presented in detail. The biopsy procedure is carefully explained, as well as the preparation of the cytology smears. Finally, the limitations and complications of the procedure are reviewed in brief, stressing the low rate of complications (below 1-2%), most of them being minor and self-limiting. Currently endosonography has strengthened its position as a diagnostic and staging method, especially after establishing the method of FNA biopsy. Thus, EUS-FNA is very useful to establish an initial tissue diagnosis of malignancy, but also to accurately stage the patients preoperatively, influencing the decision-making process and reducing the morbidity and mortality that accompanies inappropriate surgical interventions in patients with advanced cancer.