Transrectal ultrasound (TRUS) biopsy can miss 20-30% of clinically significant cancers. We evaluate an alternative approach-transperineal template-guided mapping biopsy (TTMB) in the initial and repeat biopsy setting. From January 2005 through September 2008, 373 consecutive men underwent TTMB (294 men with > or =1 prior negative biopsy and 79 men as the initial biopsy). The location of each positive biopsy core, number of positive cores, and percent involvement of each core was recorded. Cancer detection rate for the initial biopsy was 75.9%. For men with 1, 2, and > or =3 prior negative biopsies detection rates were 55.5%, 41.7%, and 34.4%, respectively. In all, 55.5% of the cancers identified were Gleason > or =7. The majority of the cancers were multifocal. There was no significant change in the number of positive cores or Gleason score as the number of prior biopsies increased. The anterior and apical aspects of the prostate were among the most common cancer locations. TTMB provides a high rate of cancer detection as initial and repeat biopsy. TTMB was particularly effective at diagnosing anterior and apical cancer. TTMB may have particular application for men considering active surveillance, with prior negative TRUS biopsies, and those considering subtotal gland or other minimally invasive treatments.