Due to lack of robust, sensitive and low cost detection strategies, Tuberculosis (TB) remains a significant global health issue. WHO reports 1.5 million deaths per year, ∼80 % cases occur in low- to middle-income countries, where resource limitations complicate the diagnosis. Robust detection of TB infection is important to contain the spread and treat disease. We developed a label-free DNA biosensor based on commercially available screen printed electrodes (SPEs) (DropSens and Zensors) that can detect TB robustly, sensitively, and specifically via DNA hybridization with its IS6110 gene marker, in purified DNA and raw sputum samples. The fabricated biosensor was morphologically characterized by scanning electron microscopy coupled with energy dispersive X-ray spectroscopy. Cyclic voltammetry and Differential Pulse Voltammetry was used for electrochemical analysis of the modified electrode. The fabricated biosensor demonstrated satisfactory selectivity for Mycobacterium tuberculosis (MTB) against Salmonella typhimurium and Escherichia coli and was able to detect MTB; the limit of detection (LOD) of 1.90 nM with R = 0.993, when analyzed over a range of concentrations of DNA (0.5-10 nM). It is being exploited to detect target MTB from clinical samples, without DNA purification. The approach is robust, sensitive, and specific, requires low sample volume and can be extended towards portable point of care diagnosis of TB.