Introduction: CNS Vital Signs (CNS VS) is a computerized neuropsychological test battery that is translated into many languages. Test-retest reliability and potential practice effects of CNS VS were evaluated. Method: Dutch healthy participants were tested with CNS VS (T0), and retested after 3 (T1) and 12 months (T2). CNS VS consists of 7 tests (i.e., Verbal Memory, Visual Memory, Finger Tapping, Symbol Digit Coding, Stroop, Shifting Attention and Continuous Performance), which cover multiple cognitive domains. Test- retest reliability was determined using Pearson’s/Spearman’s correlations. To evaluate potential practice effects, paired- sample t-tests were conducted (T0 versus T1, and T1 versus T2). Results: 159 (57% female) subjects, aged 20 to 80 years, were included. At T1 and T2, 132 and 63 subjects were retested, respectively. Low to high test-retest correlations were found for CNS VS’ tests and domains (r/ρ ranging from .17 to .89). At T1, participants performed significantly better on CNS VS’ Stroop Test, Shifting Attention Test and Symbol Digit Coding Test, resulting in significantly higher scores on the domains of Cognitive Flexibility, Processing Speed and Reaction Time, but Cohen’s d effect sizes were small. No significant differences in performance were found between T1 and T2. Discussion: The observed test-retest reliabilities varied considerably. Practice effects were present between the first and second assessment, but not thereafter. Practice effects must be considered when interpreting performance on CNS VS at follow-up assessment. Our findings are in line with previous studies that evaluated CNS VS and correspond to existing literature on conventional (paper and pencil) tests.