The effects of explanatory variables derived from a work stress model (the effort-reward imbalance model) on salivary cortisol were assessed. A multilevel analysis was used to distinguish the effects of single occasion and multiple occasion measurements of work stress and effect on cortisol. The single (or cross-sectional) factors include Effort-Reward Imbalance (ERI), need for control, negative affect, and other enduring factors (type of occupation, gender, and smoking). The multiple occasion measurements include momentary negative mood, Momentary Demand-Satisfaction Ratio (MD-SR), sleep quality, work load (workday versus day off), at work (versus not being at the workplace), and lunch. The effect of time of day on cortisol was controlled for before the effects of these variables were determined. Momentary negative mood but not trait negative affect was positively associated with ambulatory measured cortisol. The variables from the work stress model--effort, reward, need for control, and the multiple occasion measurements of demand and satisfaction--did not affect cortisol. As could be expected, time of day had an effect on cortisol, but a hypothesised interaction with momentary negative mood was not found. Additionally, the results show that the time course of cortisol differs between individuals and that the effect of sleep quality on cortisol can vary from person to person. This points to the necessity of continued efforts to single out sources of individual variability. The finding that variables derived from the effort-reward imbalance model are not related with cortisol does not support the hypothesis that ERI leads to short-term changes in cortisol, indicating no relation with hypothalamic-pituitary-adrenal (HPA) axis activity. On the other hand, the present results invite further qualification of negative affect as a potential determinant of HPA activity, at least, as far as can be deduced from cortisol measurements.