A novel colorimetric chemosensor, (Z)-3-(5-(10H-phenothiazin-10-yl)thiophen-2-yl)-2-(pyridin-2-yl)acrylonitrile (PTH-TH-PY), was synthesized through Buchwald coupling followed by Knoevenagel condensation for the detection of hypochlorite ions (OCl-). This compound demonstrates exceptional selectivity and sensitivity towards OCl- ions with a low detection limit of 0.2 nM. PTH-TH-PY's remarkable sensitivity is attributed to its effective inhibition of the internal charge transfer (ICT) mechanism. High-resolution mass spectrometry (HR-MS) and Job's plot analysis confirmed a 1:1 binding ratio between the compound and OCl-. The probe also changes from yellow to colorless upon interaction with OCl- in naked-eye. The practical utility of PTH-TH-PY was validated through TLC plate analysis and its ability to detect OCl- in various water samples. Furthermore, the compound's capacity to detect hypochlorite in live HeLa cells indicates its potential application in biological systems. These findings highlight PTH-TH-PY as a promising sensor for real-time OCl- monitoring in diverse settings, including environmental and medical contexts.