The treatment of AIDS patients has been significantly improved by the introduction of inhibitors of the HIV enzymes reverse transcriptase and protease, as well as their use in combination therapy. However, successful therapy is often complicated by the emergence of resistant viral variants as a result of mutations. Therefore, the detection of such genetic changes is of great clinical interest. In this work, an oligonucleotide microarray was developed to detect resistance-relevant mutations within the HIV protease and reverse transcriptase, which represents the relevant sequence segments. Each mutation in the HIV genome alters the pattern of fluorescence signals on the microarray during analysis, allowing for direct inference about the viral nucleic acid sequence. The developed testing system enables rapid, cost-effective, and highly specific determination of resistance to antiviral drugs. As an alternative to routinely used Sanger sequencing, it could simplify high-throughput investigations in clinical diagnostics.