Carbon monoxide (CO) is an important cause of deaths via poisoning. CO poisoning causes inhibition of O2 transport and development of tissue hypoxia, which then causes cell apoptosis. A significant indicator of cell apoptosis, soluble tumor necrosis factor-like weak inducer of apoptosis (sTWEAK) protein, is important for the stimulation of apoptosis. The primary purpose of this study is to determine whether apoptosis occurs during acute CO poisoning and to show that sTWEAK protein is an indicator of apoptosis that can be analyzed as a marker in the peripheral blood sample. The secondary aim is to determine the diagnostic and prognostic values of sTWEAK protein. The study was performed prospectively on 43 patients with CO poisoning and 30 healthy volunteer control individuals. The anamneses were taken from all patients, who also underwent physical examination. Complete blood count, biochemical markers, cardiac enzymes, and arterial blood gas measurements were analyzed. All the patients’ sTWEAK protein levels were also analyzed. The sTWEAK protein level of patients with CO poisoning was 2278 pg/mL (1197–7234), while the level of the control group was 1609 pg/mL (310–3721). The patients’ sTWEAK levels were significantly higher than the controls (area under the curve: 0.77 (0.66–0.89); p < 0.001), and the cutoff value was determined as 1895.50 pg/mL. The cutoff level had a sensitivity of 74.4%, a specificity of 76.7%, a positive predictive value of 82.0%, and a negative predictive value of 67.6%. sTWEAK is a significant indicator of apoptosis in CO poisoning that can be analyzed in the peripheral blood. However, further clinical trials are needed in terms of prognostic criteria.