Central venous oxygen saturation (ScvO2) is often used to help to guide resuscitation of critically ill patients. The standard gold technique for ScvO2 measurement is the co-oximetry (Co-oximetry_ScvO2), which is usually incorporated in most recent blood gas analyzers. However, in some hospitals, those machines are not available and only calculated ScvO2 (Calc_ScvO2) is provided. Therefore, we aimed to investigate the agreement between Co-oximetry_ScvO2 and Calc_ScvO2 in a general population of critically ill patients and septic shock patients. A total of 100 patients with a central venous catheter were included in the study. One hundred central venous blood samples were collected and analyzed using the same point-of-care blood gas analyzer, which provides both the calculated and measured ScvO2 values. Bland and Altman plot, intra-class correlation coefficient (ICC), and Cohen's Kappa coefficient were used to assess the agreement between Co-oximetry_ScvO2 and Calc_ScvO2. Multiple linear regression analysis was performed to investigate the independent explanatory variables of the difference between Co-oximetry_ScvO2 and Calc_ScvO2. In all population, Bland and Altman's analysis showed poor agreement (+4.5 -7.1, +16.1%) between the two techniques. The ICC was 0.754 (95% CI: 0.393-0.880), P< 0.001, and the Cohen's Kappa coefficient, after categorizing the two variables into two groups using a cutoff value of 70%, was 0.470 (P <0.001). In septic shock patients (49%), Bland and Altman's analysis also showed poor agreement (+5.6 -6.7 to 17.8%). The ICC was 0.720 95% CI: 0.222-0.881, and the Cohen's Kappa coefficient was 0.501 (P <0.001). Four independent variables (PcvO2, Co-oximetry_ScvO2, venous pH, and Hb) were found to be associated with the difference between the measured and calculated ScvO2 (adjusted R2 = 0.8, P<0.001), with PcvO2 being the main independent explanatory variable because of its highest absolute standardized coefficient. The area under the receiver operator characteristic curves (AUC) of PcvO2 to predict Co-oximetry_ScvO2 ≥ 70% was 0.911 95% CI: 0.837-0.959, in all patients, and 0.903 95% CI: 0.784-0.969, in septic shock patients. The best cutoff value was ≥ 36 mmHg (sensitivity, 88%; specificity, 83%), in all patients, and ≥ 35 mmHg (sensitivity, 94%; specificity, 71%) in septic shock patients. The discrepancy between the measured and calculated ScvO2 is clinically not acceptable. We do not recommend the use of calculated ScvO2 to guide resuscitation in critically ill patients. In situations where the Co-oximetry technique is not available, relying on PcvO2 to predict the measured ScvO2 value above or below 70% could be an option.