One major challenge for application of hydroxyapatite nanoparticles (nHAP) in nanomedicine is the quantitative detection method. Herein, we exploited one quantitative detection method for nHAP based on the Eu3+ fluorescent labeling via a simple chemical coprecipitation method. The trace amount of nHAP in cells and tissues can be quantitatively detected on the basis of the fluorescent quantitative determination of Eu3+ ions in nHAP crystal lattice. The lowest concentration of Eu3+ ions that can be quantitatively detected is 0.5 nM using DELFIA enhancement solution. This methodology can be broadly applicable for studying the tissue distribution and metabolization of nHAP in vivo.