Voltammetric methods hold promise for the rapid and sensitive quantification of melatonin. This study reports the direct electrochemical quantification of melatonin using carbon nanotube (CNT) fiber cross‐sections as microelectrodes. Six identical highly densified CNT fiber cross‐sections were employed to quantify melatonin in the range of 0.05–100 µM. The limit of detection and quantification were 10 and 35 nM, respectively, with a sensitivity of 0.1322 nA/µM. Interference studies with uric acid, hypoxanthine, and ascorbic acid demonstrate its performance. Real‐world application was highlighted by measuring melatonin in food, pharmaceutical, and human urine samples.