Objectives: Serotonin 2A (5-HT2A) receptor agonist binding measured with positron emission tomography (PET) may be clinically and pathophysiologically relevant and may reflect endogenous 5-HT levels in vivo. Here, we assess test-retest variability of the newly developed 5-HT2A receptor agonist PET radioligand [11C]Cimbi-36 [1] and compare brain distribution to the 5-HT2A receptor antagonist [18F]altanserin. Methods: Sixteen healthy volunteers were included in the study (mean age: 23.9±6.4 years, 6 males). All subjects were PET scanned for 120 minutes after a bolus injection with [11C]Cimbi-36. Eight subjects were re-scanned with [11C]Cimbi-36 (mean time ±SD between PET scans: 31±16 days) while other 8 subjects were PET scanned for 40 minutes with [18F]altanserin after a bolus plus constant infusion (Kbol=1.75 hours, mean time ±SD between PET scans: 133±88 days). Binding potentials (BP) were quantified with cerebellum as reference region for both radioligands. The regional non-displaceable BP (BPND) of [11C]Cimbi-36 were assessed with distribution volumes from a 2-tissue compartment model (2TCM) based on arterial input measurements and with reference tissue models, while the BP's for [18F]altanserin were assessed by radioactivities in specific brain regions and plasma assuming steady-state conditions of the radioligand. Results: The absolute differences from test to retest in [11C]Cimbi-36 BPND derived from reference tissue models in high-binding cortical regions were consistently less than 5%, indicating a higher test-retest reproducibility in measures of [11C]Cimbi-36 binding in the human brain compared to previous results with [18F]altanserin [2]. As expected, test-retest variability was slightly higher for BPND measures derived from 2TCM and in smaller brain regions. We found a highly significant correlation between regional BPNDs of [11C]Cimbi-36 and [18F]altanserin with-in subjects (mean Pearsons's r-value: 0.95±0.04) suggesting a similar regional binding of the radioligands. The highly significant correlation remained when pooling regional BPNDs from all subjects (Pearson's r=0.92, p<0.0001) and no regionally specific deviations from the correlation between the two radioligands were observed. Voxel-based analyses of mean standardized [11C]Cimbi-36 and [18F]altanserin BPND images showed that binding of [18F]altanserin relative to [11C]Cimbi-36 was higher in regions corresponding to gray matter in cerebral cortex, but lower in the white matter and in a region corresponding to the choroid plexus. Conclusions: Good reproducibility in the human brain, particular in the cortical areas, highlights the potential of [11C]Cimbi-36 as a PET radioligand in longitudinal studies assessing 5-HT2A receptor agonist binding and in studies to measure changes in binding due to fluctuations in the levels of 5-HT. [11C]Cimbi-36 and [18F]altanserin both bind with high affinity to 5-HT2A receptors, but [11C]Cimbi-36 has greater relative selectivity for 5-HT2C receptors as compared to [18F]altanserin, which is the likely explanation for the higher binding of [11C]Cimbi-36 in choroid plexus. Our results do not suggest any regional differences in binding to high-affinity versus low-affinity states of 5-HT2A receptors. References [1] Ettrup, A., da Cunha-Bang, S., McMahon, B., Lehel, S., Dyssegaard, A., et al., 2014. J. Cereb. Blood Flow Metab 34, 1188–96 [2] Haugbol, S., Pinborg, L.H., Arfan, H.M., Frokjaer., V.M., Madsen J., et al., 2007. Eur. J. Nucl. Med. Mol. Imaging 34, 910–5