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Here we report and review the investigation of bone microenvironment in vivo with harmonic generation microscopy. Excited by an infrared femtosecond laser, the second harmonic generation images can reveal the bone structures and boundaries. The third harmonic generation images can reveal the osteocytes, connecting canaliculi, and granular bone marrow cells. These imaging features can be used to observe and analyze the histology of the bone microenvironment in the future. [ABSTRACT FROM AUTHOR]

A single nanomaterial with multiple imaging contrasts and functions is highly desired for multiscale theragnosis. Herein, we demonstrate single 1–1.9 μm infrared-active FePt alloy nanoparticles (FePt NPs) offering unprecedented four-contrast-in-one molecular imaging – computed tomography (CT), magnetic resonance imaging (MRI), photoacoustic (PA) imaging, and high-order multiphoton luminescence (HOMPL) microscopy. The PA response of FePt NPs outperforms that of infrared-active gold nanorods by 3- to 5.6-fold under identical excitation fluence and particle concentrations. HOMPL (680 nm) of an isolated FePt NP renders spatial full-width-at-half-maximum values of 432 nm and 300 nm beyond the optical diffraction limit for 1230-nm and 920-nm excitation, respectively. The in vivo targeting function was successfully visualized using HOMPL, PA imaging, CT, and MRI, thereby validating FePt as a single nanomaterial system covering up to four types (Optical/PA/CT/MRI) of molecular imaging contrast, ranging from the microscopic level to whole-body scale investigation. [ABSTRACT FROM AUTHOR]