Two-photon probe for Cu²⁺ with an internal reference: quantitative estimation of Cu²⁺ in human tissues by two-photon microscopy.

Copper ions play a crucial role in living systems as cofactors of numerous metalloenzymes. To quantitatively estimate the Cu(2+) concentration in human tissue, we have developed a two-photon (TP) probe with an internal reference (ACCu2) that shows significant TP action cross-section and high selectivity for Cu(2+) and can quantitatively estimate the Cu(2+) concentration in human colon tissues by dual-color two-photon microscopy (TPM) imaging with minimum interference from other competing metal ions or pH and minimum cytotoxicity and photostability problems. The Cu(2+) concentrations in human normal colon, polyp, and colon cancer tissues were found to be 8.3 ± 0.3, 13 ± 2, and 22 ± 3 μM, respectively. This result suggests that ACCu2 may be useful for the diagnosis of human colon cancer.