It has been reported that the dimeric isoform of the enzyme pyruvate kinase M2 was overexpressed in various solid tumor cells. Hence, it was suggested that circulating levels of the so-called tumor M2-pyruvate kinase (Tu M2-PK) could be used as a tumor marker for monitoring systemic therapies of various solid tumors. We analyzed its validity as a tumor marker by comparing plasma levels of Tu M2-PK in patients with different non-malignant diseases to levels in healthy individuals and in patients with hematological diseases. Plasma levels of Tu M2-PK were measured using an ELISA assay in a total of 284 patients. The mean Tu M2-PK concentration of 32 U/mL was significantly higher in the group of patients with hematological malignancies (n = 121) (p < 0.001). However, 37% of healthy individuals (n = 63) and 44% of patients with non-malignant diseases (n = 100), especially patients with an acute inflammatory reaction (67%), were found to have elevated levels of Tu M2-PK using a cutoff level of 15 U/mL. The specificity was 59% and the sensitivity was 51%. There was no significant correlation between the prevalence of a hematological malignancy and positive Tu M2-PK result. Thus, our data imply that Tu M2-PK is not a useful tumor marker for hematological malignancies and solid tumors, as a significant number of false positive results were detected in healthy individuals and patients with non-malignant diseases.