Testicular biopsy has been widely used for the diagnosis of male infertility. Since the introduction of intracytoplasmic sperm injection (ICSI), sperm recovered from a testicular biopsy specimen can be successfully used for establishing pregnancies. Testicular spermatozoa may be recovered from testicular tissue in patients with excretory azoospermia but also in many patients with secretory azoospermia. In the latter patients spermatozoa may be recovered only after multiple excisional testicular biopsies, irrespective of follicle-stimulating hormone level, testicular size or medical history. Less invasive techniques such as percutaneous fine-needle aspiration have been introduced and may yield comparable success rates in patients with normal testicular function. The use of cryopreserved testicular spermatozoa may become an alternative to repeated surgery for obtaining testicular tissue for subsequent ICSI treatment cycles if larger series confirm the preliminary case reports. The introduction of the use of testicular spermatozoa for ICSI has raised new concerns because potentially genetically immature germ cells are being used from patients who may carry genetic defects causing their infertility problems.