In vivo multiphoton tomography and fluorescence lifetime imaging of human brain tumor tissue.

High resolution multiphoton tomography and fluorescence lifetime imaging differentiates glioma from adjacent brain in native tissue samples ex vivo. Presently, multiphoton tomography is applied in clinical dermatology and experimentally. We here present the first application of multiphoton and fluorescence lifetime imaging for in vivo imaging on humans during a neurosurgical procedure.

Comparison of morphologic criteria for actinic keratosis and squamous cell carcinoma using in vivo multiphoton tomography.

The routine diagnostic procedure of actinic keratosis (AK) and invasive squamous cell carcinoma (SCC) is a histological examination after taking a biopsy. In the past decades, non-invasive optical methods for skin examination have been developed. Patients with clinical diagnosis of AK or SCC were examined.

Multipurpose nonlinear optical imaging system for in vivo and ex vivo multimodal histology.

We report on a flexible multipurpose nonlinear microscopic imaging system based on a femtosecond excitation source and a photonic crystal fiber with multiple miniaturized time-correlated single-photon counting detectors. The system provides the simultaneous acquisition of e.g.

Cancer-cell killing by engineered Salmonella imaged by multiphoton tomography in live mice.

Our laboratory has previously developed a bacterial cancer therapy strategy by targeting tumors using engineered Salmonella typhimurium auxotrophs (S. typhimurium A1-R) that were generated to grow in viable as well as necrotic areas of tumors but not in normal tissue. The mechanism by which A1-R kills cancer cells is unknown.

Multiphoton tomography visualizes collagen fibers in the tumor microenvironment that maintain cancer-cell anchorage and shape.

Second harmonic generation (SHG) multiphoton imaging can visualize fibrillar collagen in tissues. SHG has previously shown that fibrillar collagen is altered in various types of cancer. In the present study, in vivo high resolution SHG multi-photon tomography in living mice was used to study the relationship between cancer cells and intratumor collagen fibrils.

Three-dimensional multiphoton/optical coherence tomography for diagnostic applications in dermatology.

A preliminary clinical trial using state-of-the-art multiphoton tomography (MPT) and optical coherence tomography (OCT) for three-dimensional (3D) multimodal in vivo imaging of normal skin, nevi, scars and pathologic skin lesions has been conducted. MPT enabled visualization of sub-cellular details with axial and transverse resolutions of <2 μm and <0.5 μm, respectively, from a volume of 0.

Combined in vivo multiphoton and CARS imaging of healthy and disease-affected human skin.

We present combined epi-coherent anti-Stokes Raman scattering (CARS) and multiphoton imaging with both chemical discrimination and subcellular resolution on human skin in vivo. The combination of both image modalities enables label-free imaging of the autofluorescence of endogenous fluorophores by two-photon excited fluorescence, as well as imaging of the distribution of intercellular lipids, topically applied substances and water by CARS. As an example for medical imaging, we investigated healthy and psoriasis-affected human skin with both image modalities in vivo and found indications for different lipid distributions on the cellular level.

Watching stem cells in the skin of living mice noninvasively.

We demonstrate noninvasive, high-resolution multiphoton tomography of nestin-expressing stem cells of hair follicles in living transgenic nude mice. An imaging system comprised of a compact femtosecond laser, 3D scan head mounted on a flexible mechano-optical articulated arm for simultaneous intra-tissue fluorescence and second harmonic detection (SHG) detection was used. This noninvasive method enables long-term in vivo tracking of intra-tissue stem cells in living animals.

Exploration of chromatic aberration for multiplanar imaging: proof of concept with implications for fast, efficient autofocus.

Image-based autofocus determines focus directly from the specimen (as opposed to reflective surface positioning with an offset), but sequential acquisition of a stack of images to measure resolution/sharpness and find best focus is slower than reflective positioning. Simultaneous imaging of multiple focal planes, which is also useful for 3D imaging of live cells, is faster but requires complicated optics. With color CCD cameras and white light sources commonly available, we asked if axial chromatic aberration can be utilized to acquire multiple focal planes simultaneously, and if it can be controlled through a range sufficient for practical use.

Multiphoton tissue imaging using high-NA microendoscopes and flexible scan heads for clinical studies and small animal research.

Multiphoton tomographs based on femtosecond laser and GRIN lens technology in combination with flexible scan heads have been developed for clinical high-resolution tissue imaging and small animal research. The novel tissue tomograph possesses a 0.5 m long flexible mirror arm in combination with piezodriven focusing optics and multiple single photon counting PMT detectors.