## 5 pH in Cellular Organelles
Tagging subcellular domains of Golgi is rather difficult. Many Golgi proteins reside in endoplasmic reticulum for a short period of time, thereby indicating that proteins normally cycle between the Golgi and endoplasmic reticulum [58]. Furthermore, some Golgi proteins such as TGN38/46, a _trans_-Golgi network (TGN) protein [59], and GPP130, a _cis_-Golgi protein [60], cycle between the Golgi and endosomes. Directing the pH sensor to _medial/trans_-Golgi is achieved with N-terminal linking pHluorin to \(\beta\)-1, 4-galactosidaseosyltransferase [11,12,57,61], 2,6-sialyltransferase [44], and TGN38 in a _trans_-Golgi network [59]. ManI-PpHluorin--pHluorin fused with glycine max mannosyl-oligosaccharide 1,2-alpha-mannosidase--is used to specify the _cis_-Golgi luminal pH [4]. Figure 2: The pH of individual subcellular compartments in a prototypic eukaryotic cell. We show only pH values obtained using the genetically encoded pH-sensitive FPs as indicators. The pH values are collected from different sources and are referenced in Table 4. \begin{table}
\begin{tabular}{c c c c c} \hline
**Sensor** & **Tag** & **pH** & **Organism/Cell Type** & **Instrumenta ** & **Reference** \\ \hline \multicolumn{5}{c}{Cytosol} \\ \hline E\({}^{2}\)GFP & 7.2–7.3 & CHO, U-2 OS cells & Micro & [36] \\ E\({}^{2}\)GFP & 7.3–7.5 & Rat hippocampal neuron & Micro & [48] \\ deGFP4 & 7.4–7.7 & Rat hippocampal neuron & Micro & [48] \\ GFP-F64L/S65T & 7.4 & CHO-1K cells & Micro & [11] \\ EYFP & 7.4 & HeLa cells & Micro & [12] \\ SpPrHer & 7.2 & HeLa cells & Micro & [41] \\ pHRed & 7.4 & Neuro 2A cells & Micro & [40] \\ pHERP & 7.2–7.5 & Vero, CFT1, CHO cells & Micro & [44] \\ ClopHensor & 7.3, 7.7 & WSS1, PC12, CHO cells & Micro & [45,46] \\ ClopHensor & 7.5–7.7 & CHO cells & Micro & [46] \\ (H148G/V224L) & & & & \\ PalmPalm-ClopHensor & Palmitoylation site & 7.3 & WSS1, PC12 cells & Micro & [46] \\ GFpH & C-terminus & 6.8 & COS-7, CHO-K1 cells & Micro & [43] \\ Caveolin-E\({}^{\prime}\)GFP & Caveolin-1 & 7.3 & HeLa cells & Micro & [50] \\ hCNT3-mNectarine & Cytosolic site of & 6.5–7.5 & HEK cells & Micro & [25] \\ hCNT3 & hCNT3 & 6.5–7.7 & _A. niger_ & Micro & [33,34] \\ RaVC & 7.4–7.7 & _S. cerevisiae_ FL100, S288c, & & \\ pHluorin & 6.4–7.5 & \(\sum\)1278, ORY001, BY4742 & & Spec & [52–54] \\  & & _S. cerevisiae_ H4307, H3909, & Micro & \\ pHluorin, pHluorin2 & 7.0–7.5 & \(\sum\)37473 & Flow & [31,55] \\ pHluorin & 7.2–7.3 & _Schizo. pombe_ & Micro & [49] \\ pHluorin & 7.0 & _C. glabrata_ BG14 & Spec & [51] \\ pHusion & 6.4 & _Arabidopsis_ mesophyll cell & Micro & [47] \\ pHGFP & 6.5–7.0 & _A. thaliana_ root cap & Micro & [15] \\ pHGFP & 7.0–7.3 & _A. thaliana_ meristem cells & Micro & [15] \\ PRpHluorin & 7.3 & _A. thaliana_ & Micro & [4] \\ \hline \multicolumn{5}{c}{Nucleus} \\ \hline NLS–PRpHluorin & PKKKRKV & 7.2 & _A. thaliana_ & Micro & [4] \\ pHluorin-NLS & PKKKRKV & 7.3 & _Schizo. pombe_ & Micro & [49] \\ \hline \multicolumn{5}{c}{Endoplasmic reticulum} \\ \hline pHluorin-KDEL & KDEL at C-terminus & b & Rat hippocampal neurons & Micro & [56] \\ GFP-F64L/S65T- & SEKDEL at C-terminus of FP & b & CHO-K1, LLC-PK1 cells & Micro & [11] \\ PpHluorin–HDEL & HDEL at C-terminus & 7.1 & _A. thaliana_ & Micro & [4] \\  & of FP & & & \\ \end{tabular}
\end{table} 
Table 4: Select examples of pH-sensitive FPs targeted to cytosol, nucleus, and endoplasmic reticulum. Due to the heterogeneity of secretory vesicles and their acidification during secretion, the analysis of vesicular pH presents a serious challenge (Table 6). Primarily, e-pHluorin is used for the analysis of vesicular dynamics; therefore, spatial and temporal pH changes rather than the pH of recycling vesicles are known. Synaptic vesicles maintain an acidic lumen with a resting pH value of 5.6 [13]. Therefore, instead of pHluorins, FPs with a pK\({}_{\mathrm{a}}\) of almost 6 might be better suited for pH measurements of acidic subcellular compartments, including maturing endosomes.