Determination of zinc using carbonic anhydrase-based fluorescence biosensors.

This chapter summarizes the use of carbonic anhydrase (CA)-based fluorescent indicators to determine free zinc in solution, in cells, and in subcellular organelles. Expression (both in situ and in vitro) and preparation of CA-based indicators are described, together with techniques of their use, and procedures to minimize contamination. Recipes for zinc buffers are supplied.

DsRed as a highly sensitive, selective, and reversible fluorescence-based biosensor for both Cu(+) and Cu(2+) ions.

The wild type form of Red fluorescent protein (DsRed), an intrinsically fluorescent protein found in tropical corals, is found to be highly selective, reversible and sensitive for both Cu(+) and Cu(2+), with a nanomolar detection limit. The selectivity towards these ions is retained even in the presence of other heavy metal ions. The K(d) values for monovalent and divalent copper, based on single binding isotherms, are 450 and 540 nM, respectively.