Capillary electrophoresis-SELEX (CE-SELEX) was used to select ssDNA aptamers with affinity for HIV reverse transcriptase (HIVRT). A library of ssDNA was incubated with HIVRT. Sequences bound to HIVRT were isolated using CE, PCR amplified, and purified, yielding an enriched ssDNA pool suitable for further rounds of selection. Aptamers with dissociation constants as low as 180 pM were isolated after four rounds of selection. This is the first report of aptamers isolated by CE-SELEX with higher affinity than those obtained for the same target using conventional selection techniques. No sequence motifs were identified in the 27 clones sequenced, suggesting that there are many sequences that can bind HIVRT with low picomolar dissociation constants.