A highly sensitive ratiometric fluorescence pH probe for extracellular pH in mouse brain slice

High-sensitive detection of extracellular pH is prerequisite for finding the abnormality of several serious diseases at early stage and clarifying their pathogenesis. However, the present fluorescence pH probes were deficient in sensitivity to accurately detect extracellular pH. Here, we fabricated a high-sensitive ratiometric fluorescence pH probe, SiRho-F-pH, through the click reaction between a Si-rhodamine and a fluorescein, which had the opposite pH-responses. Upon 450 nm light excitation on the spectrometer, the ratio R_680/520 of fluorescence intensities of SiRho-F-pH at 680 nm and 520 nm changed by 1065-fold in the pH range of 4.57-9.33. While, SiRho-F-pH possessed the excellently reversible pH-response, high pH-selectivity, favorable biocompatibility, and ability to target the detection of extracellular pH. The microscope imaging at fixed cells stained with SiRho-F-pH showed that the ratio R/G of fluorescence from red and green channels changed by a notable 4858-fold in the pH range of 5.00-8.00 under 488 nm laser excitation. The mouse brain slice imaging further demonstrated that SiRho-F-pH could clearly reveal tiny changes of extracellular pH in the pH range of 6.61-6.71 with micrometer spatial resolution. The ratiometric fluorescence mode of SiRho-F-pH could be used to detect the extracellular pH with high sensitivity and spatial-resolution.