## 1 Introduction  
The regulation and homeostasis of pH within a cell and its subcellular compartments are crucial for the viability of any living cell, from the simplest prokaryotes to complex multicellular organisms. Stringent pH requirements are necessary for efficient metabolism, protein stability, ion channel activity, membrane trafficking, protein sorting, and proteolytic processing of proteins. Cellular compartments are protected from rapid, local changes in pH by intrinsic buffering capacity that is provided by various intracellular weak acids and bases. A more dynamic network for controllingintracellular pH (pH\({}_{i}\)) is pH homeostasis, which is composed of an ensemble of well-coordinated ion carriers, such as ion permeable channels, transporters, and pumps [1; 2; 3; 4].  
An important step forward in the understanding pH homeostasis has been achieved with the development of genetically encoded fluorescent pH-sensitive sensors, which provide a means for spatial and temporal imaging of pH dynamics. Fluorescent proteins (FPs) as intrinsic intracellular reporters have many advantages over conventional fluorescent dyes in live cell imaging. They are genetically encoded, therefore, no loading of dye is necessary, thereby enabling noninvasive imaging. The FPs can be precisely targeted to almost any organelle, compartment, or tissue. Moreover, FPs have been designed to respond to a greater variety of biological events and signals than conventional dyes. The sensors based on FPs have been used in living cells to report cellular concentrations of ions, second messengers cAMP and inositol phosphates, ATP, redox potential, reactive oxygen species, enzyme activities, and pH (review [5]).  
The use of FPs began with the identification and subsequent characterization of wild-type green fluorescent protein (wtGFP) from the jellyfish _Aequorea victoria_[5; 6]. Within the \(\beta\)-barrel structure of GFP, three consecutive amino acids S65, Y66, G67 form by post-translational cyclization a chromophore that, by itself, is not fluorescent. Amino acids Q69, Q94, R96, H148, T203, S205, and E222 that are placed around the chromophore are necessary for GFP fluorescence [7]. These residues function as proton donors and acceptors and define spectral and photophysical properties of GFPs [8]. [PERSON] _et al._[9] provide a detailed review of the physical and chemical characteristics of the chromophores of different GFP mutants. Briefly, the optical properties of protonated-neutral and deprotonated-anionic states differ among FPs. Among other parameters, pH also affects the equilibrium between protonated and deprotonated forms. Some FPs show pH-dependent absorption/  
Figure 1: **(A)** A two state model of pH dependent ground states of wtGFP. Four molecules of water and side-chains of amino acids depicted in grey are involved in hydrogen-bonding with the chromophore. (**B**) Chromophore formation involves cyclization, imidazolinone ring system formation, dehydration and oxidation.  
fluorescence spectra in the physiological pH range and are, therefore, used as pH sensors for live cell imaging (Figure 1).  
Further, pH-sensitive FPs are divided in two groups: (i) intensity-based sensors and (ii) ratiometric pH-sensitive proteins with dual excitation and/or emission spectra. There is also a third group of pH-sensitive proteins--chimeric proteins, which are concatenated pH-sensitive FPs with another fluorescent or luminescent protein that gain novel pH-dependent spectral properties through fusion. Some of the most frequently used pH sensors are described below.