Gene transfer vectors based on the replication-defective human parvovirus, adeno-associated virus type 2 (AAV-2), are viable candidates for in vivo and ex vivo human use. However, widespread testing of AAV vectors has been limited by difficulties in generating pure, high-titer vector stocks that are fully characterized. To address these issues, we have developed a single-step purification scheme using heparin affinity chromatography. Recovery from the crude lysate starting material exceeds 70%, and the end product rAAV vector is highly purified and appears to be free of adenovirus and cellular contaminates. Importantly, purified vectors retain predicted in vivo biologic activity. Concurrently, we have developed simple and rapid approaches for vector quantification using real-time PCR. These new methods, combined with the use of stable producer cell lines for rAAV production, make the commercial production of rAAV vectors for human use truly viable and pragmatic.