In modern forensic genetics, the analysis of DNA mixtures is a challenge because of the obstacles found in defining the number of contributors, especially when they exceed two. The goal of the present study was to find the most reliable method of detection for profiling the two-male and female-male mixtures obtained. Bloodstains, saliva, and body tissues from various crime scenes were obtained as samples. All samples for DNA profiling have been subjected to DNA extraction, amplification, separation, and detection. Results revealed that DNA profiling showed two-male and female-male mixtures. Results also showed the presence of a major and a minor male contributor in each genetic locus according to peak heights. Differential extraction provided a reliable method for separating female and male contributors from each other in the case of the female-male mixture. The profiling of the male donor in the female-male mixture was furtherly verified using the Y-STR profiling process. Eventually, we can infer that a good approach to identifying the two-male mixture is autosomal STR interpretation centered on major and minor differentiation. In addition, differential separation and Y-STR profiling proved to be the best processes for profiling mixed samples comprising female-male contributors as only the male profile was identified.