pH, CO32−, and Ca2+ liquid ion exchange (LIX) microsensors were prepared as described previously (4346), with the exception that the green glass tubing was replaced by glass capillary tubes (borosilicate; inner diameter, 1.2 mm; outer diameter, 1.5 mm; Hilgenberg), which were pulled on a DMZ Universal puller (Zeitz-Instruments) into micropipettes with tip diameters of 1 μm. In brief, the micropipette tips were silanized (43), backfilled with electrolyte {for H+: 300 mM KCl, 50 mM NaPO4, pH 7; for CO32−: Na2B4O7 × 10H2O (19.1 g liter−1) + 200 μM CaCO3 adjusted with 0.1 N HCl and NaOH to pH 9 [see (44)]; and for Ca2+: 200 mM KCl, 2 mM CaCl2 × 2H2O}, and broke open under the microscope to tip diameters of 2 to 3 μm. Tips were then front-filled in two steps with H+, CO32−, or Ca2+ ionophore, respectively (H+ ionophore III, CO32− ionophore VII, and Ca ionophore II; Sigma-Aldrich), as described previously (44). The micropipette tips were then glued to the rest of the glass housing, and microsensors were left to dry for 2 to 6 hours before measurements.

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