An advanced detection assay is important for clinical diagnosis of human immunodeficiency virus (HIV) infection in asymptomatic individuals. The first step in timely treatment and management of the infection is the early detection of HIV. In this study, we determined the prevalence of HIV in Islamabad for the first time using a chemiluminescent microparticle immunoassay (CMIA), which is an advanced serological diagnostic technique. A total of 3659 samples were obtained from the general public in Islamabad, Pakistan, and 85 of them were found to be positive for HIV infection (2.32% prevalence). Of the positive subjects, 78.82% (67/85) were male, 10.6% (9/85) were female, and 10.6% (9/85) were of unidentified gender. The results revealed a significant relationship between age groups and HIV status. The age group of 21- to 30-year-olds was found to have the highest rate of HIV infection, and the rate of HIV infection in males was higher than in females.