Objective: To determine the prevalence of Y chromosome microdeletions in Pakistani idiopathic infertile men, using multiplex polymerase chain reaction. Patients and Methods: A case control study was conducted on the infertile male patients attending OPD’s of Aziz Medical Hospital and National Center for Fertility Control, Jinnah Post Graduate Medical Center (JPMC), Karachi. A total of 220 primary infertile men, of which 150 (68.2%) had azoospermia, 40 (18.2%) had severe oligozoospermia and 30 (13.6%) had oligozoospermia and 220 fertile men as control group were studied. Six sequence-tagged sites: sY84 and sY86 for AZFa, sY127 and sY134 for AZFb, and sY254 and sY255 for AZFc were used for amplification of the azoospermia factor region of Y chromosome according to the recommendations of European Academy of Andrology and the European Quality Monitoring Network Group. Results: Yq microdeletions were found in (12) 5.45% cases, while none in the control group (p =<0.007). All the microdeletions were found in azoospermics 12/150 (8.0%). Among patients with microdeletions, the AZFa region was found to be deleted in 1 (8.33%), AZFb in 2 (16.67%), AZFc in 6 (50%), AZFb+c in 2 (16.67%) and complete AZF deletions in 1 (8.33%) patients. Identification of Y chromosome microdeletions in Pakistani infertile males was found to have significant diagnostic and prognostic value and provides useful information for genetic counseling in patients choosing assisted reproductive treatment