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ABSTRACT Neuroticism is widely used as an explanatory concept in etiological research of psychopathology. To clarify what neuroticism actually represents, we investigated the phenotypic and genetic relationship between neuroticism and the morning cortisol secretion. In the current classic twin study, 125 female twin pairs (74 monozygotic and 51 dizygotic pairs) participated. For each participant, 4 different neuroticism scores were available to calculate a neuroticism composite score that was used in the statistical analyses. The morning cortisol secretion was assessed by 4 salivary samples in the 1st hour after awakening. Significant genetic influences for the neuroticism composite score (55%), and each of the 4 cortisol samples (52%–69%) were found. There was no phenotypic or genotypic relationship between neuroticism and morning cortisol secretion. Although neuroticism and cortisol were both heritable traits, they did not share any genetic influences.