Background and Purpose: While penumbra assessment has become an important part of the clinical decision making for acute stroke patients, there is a lack of studies measuring the reliability and reproducibility of defined assessment techniques in the clinical setting. Our aim was to determine reliability and reproducibility of different types of three-dimensional penumbra assessment methods in stroke patients who underwent whole brain CT perfusion imaging (WB-CTP). Materials and Methods: We included 29 patients with a confirmed MCA infarction who underwent initial WB-CTP with a scan coverage of 100 mm in the z-axis. Two blinded and experienced readers assessed the flow-volume-mismatch twice and in two quantitative ways: Performing a volumetric mismatch analysis using OsiriX imaging software (MMVOL) and visual estimation of mismatch (MMEST). Complementarily, the semiquantitative Alberta Stroke Programme Early CT Score for CT perfusion was used to define mismatch (MMASPECTS). A favorable penumbral pattern was defined by a mismatch of >= 30% in combination with a cerebral blood flow deficit of <= 90 ml and an MMASPECTS score of >= 1, respectively. Inter-and intrareader agreement was determined by Kappa-values and ICCs. Results: Overall, MMVOL showed considerably higher inter-/intrareader agreement (ICCs: 0.751/0.843) compared to MMEST (0.292/0.749). In the subgroup of large (>= 50 mL) perfusion deficits, inter-and intrareader agreement of MMVOL was excellent (ICCs: 0.961/0.942), while MMEST interreader agreement was poor (0.415) and intrareader agreement was good (0.919). With respect to penumbra classification, MMVOL showed the highest agreement (interreader agreement: 25 agreements/4 non-agreements/kappa: 0.595;intrareader agreement 27/2/0.833), followed by MMEST (22/7/0.471;23/6/0.577), and MMASPECTS (18/11/0.133;21/8/0.340). Conclusion: The evaluated approach of volumetric mismatch assessment is superior to pure visual and ASPECTS penumbra pattern assessment in WB-CTP and helps to precisely judge the extent of 3-dimensional mismatch in acute stroke patients.