Hypochlorous acid (HOCl) is one of the reactive oxygen species (ROS) generated by the enzyme myeloperoxidase (MPO) which has both protecting as well as deleterious effects in the body. While HOCl and MPO is essential for the pathogen clearance mechanisms like Phagocytosis, ETosis etc. it is also implicated in inflammatory diseases like cardiovascular diseases, multiple sclerosis etc. and the resultant organ failure. Current small molecule based fluorescent sensing for HOCl are limited by a use of single wavelength, hence the detected HOCl levels will reflect the availability of the sensor. Here using a DNA-based platform I have developed a combination reporter, cHOClate, which simultaneously images HOCl and pH quantitatively. Phagosome targeted cHOClate could successfully map phagosomal production HOCl, as a function of phagosome maturation in live phagocytes. In macrophages phagosomal acidification was gradual and HOCl was released in a burst after acidification was achieved; while neutrophil phagosomes stay near neutral where HOCl burst happens within 5-10 minutes. Further, MPO activity in innate immune cells follows the order of macrophages< monocytes< neutrophils which is in concordance with the reported levels of MPO in these cells. This platform can be extended to simultaneous sensing of other ROS, small molecules and ions during phagosome maturation and impact of each of these species in pathogen clearance. Further cHOClate can be used to reveal the activity of MPO in other subcellular locations.




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