We present a microchip for isolation of aptamers that bind to target ligands at prespecified temperatures. The device uses integrated resistive heaters and sensors to control the temperatures of (poly)dimethylsiloxane (PDMS) microchambers for temperature-specific selection and bead-based amplification of aptamers. Aptamers are isolated from a randomized DNA library at specified temperatures, and amplified onto microbeads using bead-based polymerase chain reaction (PCR). As a proof of concept, the device was used to isolate aptamers for human immunoglobulin E (IgE), with the enriched pool of candidate aptamers exhibiting a much higher and temperature-dependent affinity for the target protein. The procedure was performed with significantly less reagent use in a much shorter time period (4 hours) than with conventional devices (up to 2 days).