Abstract Sulfadimethoxine (SDM) and ormetoprim (OMP) residues were determined in catfish fillets after treatment with Romet. A liquid chromatographic (LC) method capable of simultaneously detecting SDM and OMP at 0.05–40 ppm was used. The recoveries were 92% for SDM and 108% for OMP. The intra- assay variabilities were determined for SDM and OMP at fortification levels of 0.05–40 ppm, and a coefficient of variation (CV) of less than 10% was achieved at all levels. The interassay variations were determined at fortification levels of 0.1 ppm (CVs: SDM, 6.4%; OMP, 4.9%) and 1.0 ppm (CVs: SDM, 2.8%; OMP, 3.5%). SDM and OMP residues in catfish fillets were rapidly depleted after treatment with Romet. By day 2 posttreatment, SDM and OMP were essentially nondetectable. The use of an enzyme-linked immunoassay (ELISA)-based rapid diagnostic test to determine SDM and OMP in catfish fillets was evaluated. An assay procedure to adapt the test for residues in tissue samples was developed. SDM and OMP were extracted from ground catfish fillet with methanol–water (80 + 20, v/v); the extract was diluted with buffer and filtered, and the filtrate was then tested with an EZ Screen test card. The effectiveness of the rapid test assay to detect SDM residues at levels above the U.S. Food and Drug Administration tolerance (0.1 ppm) was verified with fillets from Romet-treated catfish. All samples with residues at >0.1 ppm were identified correctly by the test. The results indicate that the diagnostic test could be used as a rapid method for monitoring Romet residues in catfish.