In this work, with the aim to develop effective molecular probes and investigate the structure-reactivity correlation, we designed a short series of phenothiazine based fluorescent probes for detecting ClO with differing electron push-pull groups. Sensing experiment results and single crystal X-ray analysis with the aid of TD-DFT calculation revealed that substituting groups with increasing electron-withdrawing ability could increase the dihedral angle of phenothiazine moiety and reduce the gap energy of probes, which led to enhanced reactivity towards ClO. Both PT1 and PT2 showed two-color switching upon detection of ClO. PT1 with strong electron donating group thiophene showed fluorescence color switch from salmon to blue. PT2 with medium electron donating/accepting group benzothiazole showed a fluorescence color switch from red to green. However, both PT1 and PT2 almost had no response to ONOO. Through introducing strong electron-withdrawing ketone combined with a cyano group, PT3 showed a cyan emission upon detection of ClO and weak red emission upon detection of ONOO. HRMS and 1H NMR results confirmed that PT1 and PT2 had the same sensing mode, in which the divalent sulfur of phenothiazine could be oxidized to sulfoxide by ClO. When reacted with ClO, PT3 experienced two-step reactions. It was firstly oxidized into sulphone structure by ClO and then transformed into sulfoxide phenothiazine aldehyde. When encountered ONOO, PT3 changed into aldehyde structure and some non-fluorescence byproducts. Owing to their special selectively and high sensitivity, PT1 and PT2 were applied to image the endogenous ClO in macrophage cells and zebrafish larvae. This study was expected to provide a useful guideline for probe design for various applications.