Hydroxyapatite nanoparticle based fluorometric determination and imaging of cysteine and homocysteine in living cells

Fluorescent hydroxyapatite nanoparticles (HAP-NPs) were prepared by reacting calcium ion with phosphate in the presence of Eu(III) ion. The HAP-NPs display large Stokes' shift and two strong fluorescence emissions with peaks at 590 nm and 615 nm when excited at 250 nm. The HAP-NPs also have good photostability and water solubility. The HAP-NPs combined with Cu(II) were applied to fluorometric determination of cysteine and homocysteine in biological samples and in living cells. In this detection scheme, the fluorescence of HAP-NPs is initially quenched by Cu(II). The addition of biothiols results in the formation of Cu(II)-thiol complexes and leads to fluorescence recovery. The assay allows cysteine to be detected with a 110 nM detection limit, and homocysteine with a 160 nM detection limit. The assay was successfully applied to the analysis of cysteine in spiked human serum samples and to imaging of cysteine in HeLa cells, and this demonstrates its potential for clinical testing and in biomedical research.