Fluorescent Potassium Indicators
The importance of the potassium ion (K+) is coupled to the sodium ion (Na+), because the cell expends a major part of its metabolic energy maintaining the concentrations of Na+ and K+ within the cell. Intracellular concentration ranges are 10-40 mM for Na+ and 120-400 mM for K+. Extracellular concentration ranges are 4-40 mM for K+ and 120-400 mM for Na+.
In the absence of a K+ indicator, efforts have been directed to using indirect techniques to measure or detect K+, including
- analogs like thallium or cesium to monitor K+ fluxes;
- the pH indicator BCECF AM and the ionophore nigericin in flow cytometry studies;
- combinations of ion selective carriers;
- ion-channel mediated K+ fluxes with membrane potential changes registered by voltage sensitive dyes; and
- fiber-optic sensors for K+ with pH sensitive dyes.
These alternate techniques were necessary because the previously reported fluorescent potassium indicator PBFI9 requires UV excitation and suffers from poor loading and poor brightness. The new TEFLabs indicators, Asante Potassium Green 1 and 2 (APG-1 and APG-2) successfully address these problems. The properties of the K+ indicators are shown in Table 3.1.
Table 3.1. Properties of Potassium Indicators
c The titrations to determine Kd were performed in 140 mM TMACl, 10 mM MOPS, pH 7.1 with the addition of sodium chloride. No corrections were made for increasing ionic strength or quenching by chloride.
d Once the AM ester form permeates the cell membrane, non-specific esterases hydrolyze the AM esters, taking the indicator to its active salt form.
e The excitation maximum is 517 nm; 488 nm excitation gives 40% of the maximum.
Table 3.2 compares PBFI and Asante Potassium Green.