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A few probes have been devised to measure intracellular enzyme activities by fluorescent imaging, but the study of the regulation of enzyme activity via gasotransmitters in situ remains a long\u2010standing challenge. Herein, we report a three\u2010channel imaging correlation by a single dual\u2010reactive fluorescent probe to measure the dependence of phosphatase activity on the H<jats:sub>2<\/jats:sub>S level in cells. The two sites of the probe reactive to H<jats:sub>2<\/jats:sub>S and phosphatase individually produce blue and green fluorescent responses, respectively, and resonance energy transfer can be triggered by their coexistence. Fluorescent analysis based on the three\u2010channel imaging correlation shows that cells have an ideal level of H<jats:sub>2<\/jats:sub>S to promote phosphatase activity up to its maximum. Significantly, a slight deviation from this H<jats:sub>2<\/jats:sub>S level leads to a sharp decrease of phosphatase activity. The discovery further strengthens our understanding of the importance of H<jats:sub>2<\/jats:sub>S in cellular signaling and in various human diseases.<\/jats:p>","DOI":"10.1002\/anie.201811391","type":"journal-article","created":{"date-parts":[[2018,12,27]],"date-time":"2018-12-27T10:59:31Z","timestamp":1545908371000},"page":"2261-2265","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":59,"title":["Gasotransmitter Regulation of Phosphatase Activity in Live Cells Studied by Three\u2010Channel Imaging Correlation"],"prefix":"10.1002","volume":"58","author":[{"given":"Pan","family":"Ou","sequence":"first","affiliation":[{"name":"School of Chemistry and Chemical Engineering, and Institute of Physical Science and Information Technology Anhui University  Hefei Anhui 230601 China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ruilong","family":"Zhang","sequence":"additional","affiliation":[{"name":"School of 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