Tryptophan synthase catalyzes the synthesis of a wide array of noncanonical amino acids and is an attractive target for directed evolution. Droplet microfluidics offers an ultrahigh throughput approach to directed evolution (up to 107 experiments per day), enabling the search for biocatalysts in wider regions of sequence space with reagent consumption minimized to the picoliter volume (per library member). While the majority of screening campaigns in this format on record relied on an optically active reaction product, a new assay is needed for tryptophan synthase. Tryptophan is not fluorogenic in the visible light spectrum and thus falls outside the scope of conventional droplet microfluidic readouts, which are incompatible with UV light detection at high throughput. Here, we engineer a tryptophan DNA aptamer into a sensor to quantitatively report on tryptophan production in droplets. The utility of the sensor was validated by identifying five-fold improved tryptophan synthases from ∼100,000 protein variants. More generally, this work establishes the use of DNA-aptamer sensors with a fluorogenic read-out in widening the scope of droplet microfluidic evolution.