The incidence of rape has increased, especially in metropolitan areas, such as the city of São Paulo. In Brazil, studies about it have shown that the majority of this type of crime is committed by the relatives and persons close to the victim. This has made the crime more difficult to be denounced, as only 10% of the cases are reported to competent police authorities. Usually, cytological exams are carried out in sex crime investigations. The difficulty in showing the presence of spermatozoa is frequent, but it does not exclude the presence of male DNA. The absence of spermatozoa in material collected from rape victims can be due to several factors, including the fact that the agressor suffers from azoospermia. This condition can be the result of a successful vasectomy. As the majority of DNA in the ejaculation sample is from spermatozoa, there is much less DNA to be analyzed. This study presents the application of Y-STRs (DYS19, DYS389I, DYS389II, DYS390, DYS391, DYS392, and DYS393) in DNA analysis of sperm samples from 105 vasectomized men. The study demonstrated a great variation in DNA concentration. DNA extraction and amplification was possible in all sperm samples even in the absence of spermatozoa. The same profile was observed, for each individual, from DNA extracted from blood, pre- and postvasectomy semen samples. The use of markers specific for Y chromosome in sex crime cases, especially in the absence of spermatozoa, is very important, mainly because in most situations there is a small quantity of the agressor's DNA in the medium and a large quantity of the victim's DNA.