Background. The aim of the present study was to examine sources of variation for serum cystatin C in a healthy Greek population. Methods. Cystatin C together with basic clinical chemistry tests was measured in a total of 490 adults (46 ± 16 yrs, 40% males) who underwent an annual health check. Demographic, anthropometric, and lifestyle characteristics were recorded. Results. Higher values of cystatin C were observed among males (P = .04), participants aged over 65 years (P < .001), current smokers (P = .001) and overweight/obese participants (P = .03). On the contrary, alcohol consumption and physical activity seemed to have no influence on cystatin C levels (P = .61; P = .95, resp.). Conclusions. In interpreting serum cystatin C values in a healthy adult population, age, gender, Body Mass Index, and cigarette smoking need to be considered, and determination of reference ranges among distinct subpopulations seem to be prudent.