Emerging advances in iron oxide nanoparticles exploit their high magnetization for various applications, such as catalysis, bioseparation, hyperthermia, and magnetic resonance imaging (MRI). In contrast to the excellent magnetic performance, their upconverted photoluminescence have not been thoroughly explored, thus limiting their development as a tool in photomedicine. In this work, we develop a seed/growth-inspired synthesis combined with primary mineralization and a ligand-assisted secondary growth strategy to prepare mesostructured α-FeOOH nanorods (NRs). Because α-FeOOH rods are all iron-based composites, they exhibit low cytotoxicity towards cells. Surprisingly, these mesoporous α-FeOOH mesostructures display strong third harmonic generation (THG) signals under near-infrared excited wavelength at 1230 nm. They exhibited a much stronger THG intensity compared to naked α-FeOOH NRs. Using these unique nonlinear optical properties, we demonstrate that α-FeOOH rods can serve as contrast agents in THG microscopy for the cell tracking as well as angiography in vivo. Vessel walls can be revealed after the clearance of particles. Our results provide a new strategy of material synthesis for obtaining high THG imaging contrast.
Nanomedicine can kill tumor cells or block the growth and spread of cancers by maximizing drug accumulation in tumor tissues. How much drug will selectively accumulate in tumor tissues depends on physical parameters such as vessel permeability in tumor microenvironments. On the other hand, by properly choosing the size of nanomedicine, the accumulation of drugs in normal tissues will be reduced. In this study, we measured the permeability rate of vessels in different strain of mice in vivo with two-photon fluorescent angiography. We used FITC / TRITC / TEXAS RED labeled dextran to investigate size-dependent permeation from vessels in normal tissues. We choose nanoparticles of 40kDa, 70 kDa, 150 kDa and 2000 kDa, and selected three strains of wild-type Balb/C, ICR, C2J mice. We found , even at the same size, the vascular permeability rate of dextran still vary with the dye-conjugates and the strain of mice. One of our results shows that ICR mice have the biggest vessel hole in normal tissues. Choosing wrong nanoparticle size will cause their leaking in normal tissues. This study will provide a new threshold and guideline to reduce the accumulation of nanomedicine in normal tissues.
Without a labeling, we demonstrated that lipid granules in leukocytes have distinctive third harmonic generation (THG) contrast. Excited by a 1230nm femtosecond laser, THG signals were generated at a significantly higher level in neutrophils than other mononuclear cells, whereas signals in agranular lymphocytes were one order smaller. These characteristic THG features can also be observed in vivo to trace the newly recruited leukocytes following lipopolysaccharide (LPS) challenge. Furthermore, using video-rate THG microscopy, we also captured images of blood cells in human capillaries. Quite different from red-blood-cells, every now and then, round and granule rich blood cells with strong THG contrast appeared in circulation. The corresponding volume densities in blood, evaluated from their frequencies of appearance and the velocity of circulation, fall within the physiological range of human white blood cell counts. These results suggested that labeling-free THG imaging may provide timely tracing of leukocyte movement and hematology inspection without disturbing the normal cellular or physiological status.
We have demonstrated a straightforward and noninvasive method to identify the distribution of grana
and starch within an intact leaf. Grana and starch are the major functional structures for
photosynthesis and energy storage of plant, respectively. Both exhibit highly ordered molecular
structures and appear as micrometer-sized granules inside chloroplasts. In order to distinguish grana
and starch, we used multiphoton microscopy, with simultaneous acquisition of two photon fluorescence
(2PF) and second harmonic generation (SHG) signals. Consequently, SHG is found on both grana and
starch while 2PF from chlorophyll indicates the identity of grana.
The understanding of the interaction between tumors and surrounding microenvironment in vivo is an important first step
and basis for pathway-targeting cancer therapy. To in vivo observe the dynamic development of tumor cells and validate
the efficacy of therapy in microscopic scales, people commonly performed multi-photon fluorescence microscopy
through an invasive window chamber setup. However, under such system, the cancer cells can't be identified and
long-term tracked without a fluorescence labeling. Exploiting the intrinsic third harmonic generation (THG) and
two-photon fluorescence (2PF) contrasts of melanin, we demonstrated in vivo identification of melanoma and tracked its
development without labeling. It was achieved with a least invasive femtosecond Cr:forsterite laser and a laser scanning
nonlinear microscopy system with 3D sub-micron spatial resolution. Combined with molecular probes or reporters, we
anticipate thus developed platform a powerful tool to reveal molecular insights of tumor microenvironments, enhance
our understanding of tumor biology, and trigger new therapeutic approaches.
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