Protriptyline, a tricyclic antidepressant, was converted to its heptafluorobutyramide using heptafluorobutyrylimidazole as the acylating agent. Conditions for the gas chromatography of the derivative and its quantitation by an electron capture detector were established. The smallest detectable amount of the protriptyline derivative was 0.05 nanogram. Isolation and purification procedures were devised which permit the determination of the drug in biological materials without interference from reagents, drug metabolites or naturally occurring substances. Protriptyline extracted from plasma, erythrocytes, urine and tissues was determined in concentrations as low as 10 nanograms/ml.