An assay of atenolol in human serum is described using a 3 μm silica column with a 60:40 v/v 6.25 mM sodium dihydrogen phosphate pH 3.0-acetonitrile mobile phase at 1 mL/min; the ultraviolet detector was set at 254 nm. Minimum detectability of the drug was estimated to be 20 ng/mL (S/N = 2). The method is linear in the 100–1000 ng/mL range (r = 0.9992, n = 9). An ethylsilane solid phase extraction column was used for sample cleanup and recovery of atenolol from serum was 103 ± 4% (n = 3). Retention times for atenolol and metoprolol (internal standard) were 6.6 and 7.5 min, respectively, and typical theoretical plates were 17,583 and 22,680, respectively. Resolution between internal standard and atenolol peaks was calculated to be 4.5. A typical analysis was completed in less than 45 min. The good sensitivity and reproducibility of the method should render it useful for the assay of atenolol levels in serum.