(465f) Second Window Near Infrared Fluorescent Imaging for Deep Tissue Animal Imaging

Biomedical imaging is an integral part of clinical decision-making during screening, diagnosis, staging, therapy planning and guidance, treatment and real-time monitoring of patient response, because of its ability to provide morphological, structural, metabolic and functional information at various spatio-temporal scales of interest, while being a minimally invasive and highly targeted source of physiological evidence. The main clinical challenge, however, is to be able to image biological features with sufficient sensitivity for detection at the cellular level (in a complex tissue environment at deep penetration), which could then be used to identify and treat small tumor masses  before the angiogenic switch growth phase, which is below the threshold of detection of most current imaging technologies. There are few main imaging modalities available clinically, classified according to the image contrast mechanism: X-ray (2D film imaging and computed tomography, CT), positron emission tomography (PET), single photon emission computed tomography (SPECT), magnetic resonance imaging (MRI), ultrasound (US), intravital microscopy (confocal, multiphoton), optical imaging, or some combination of these. Despite the significant advances in both the imaging instrumentation and algorithms for image processing, most of the aforementioned techniques suffer from poor sensitivity and/or resolution and/or depth of focus in the three spatial dimensions (3D). The most promising technique for high-resolution deep-tissue whole body imaging, using relatively safe molecular probes and excitation sources, at a reasonably low cost, appears to be optical imaging.

            In recent years, there has been tremendous interest in the exploration of optical imaging in vivo, due to the development of near-infrared fluorescent probes, effective targeting agents, and custom-built imagers. Particularly, NIR light has been theoretically predicted to penetrate through ~ 10 cm of human tissue, with simulations suggesting a signal-to-noise (S/N) improvement of over 100-fold by imaging in the second near-infrared window (NIR-II: 900 - 1,400 nm) compared to the first window (NIR-I: 650 - 900 nm). Despite the recent development of various NIR-II fluorescent probes and custom built imagers based on InGaAs detectors, there are three significant challenges to imaging in the NIR-II wavelength domain with InGaAs detectors. The first challenge is the limited selection of fluorescent probes which emit in the NIR-II range favorable for medical imaging. Similarly to NIR-I or visible fluorescent imaging, only part of the fluorescent signal is utilized for contrast imaging to avoid spectral overlap with autofluorescence. The second challenge is to maximize the S/N ratio for high-sensitivity deep-tissue imaging. Due to the intrinsic low SNR of InGaAs detectors compared to Si detectors, strategies are required to depress SNR of InGaAs detectors by either imaging methodologies or data processing algorithms. The third challenge is diffuse light scattering by heterogeneous turbid biological media, imposing trade-off between depth and resolution.

In this work, we designed an optical system in NIR-II window that combines both hyperspectral and hyperdiffuse measuring. HSI resolves the first two challenges of SNR by providing information in frequency domain, and allows novel type of investigation (including pixel-wise spectral analysis and 3-D reconstruction) as well as improves result confidence. HDI resolves the third challenge, by not only excluding the emission scattering in the processed results, also presenting pixel-wise diffuse scattering information for contrast imaging and 3-D reconstruction. The hyperspectral and diffuse imaging system (900-1700 nm) has the capability to distinguish the optical signatures of the primary pump laser, background, various types of tissue autofluorescence, reporter fluorescence, as well as the diffuse scattering effect of the fluorescence signal upon transport through heterogeneous turbid optical media. Further by applying our imaging system to various tissue and whole-animal, we demonstrated that it is capable of (0) no background or autofluorescence information is required to be collected beforehand, and instead, these background and aufluorescence information is proved to be useful for both diagnostic and localization reasons; (1) detecting 1 mm sized particles up to 9 cm depth through a breast-tissue mimic phantom; (2) identifying spectral and diffuse signature of various tissues; (3) reconstructing 3-D fluorescent images based on newly interpreted information from HSC and HDC; (4) identifying important organs using label-free reflection mode hyperspectral imaging for feature location and registration. This study opens up exciting new possibilities for clinical translation of NIR-II imaging as a viable platform theranostic technology; for early diagnostics, as a real-time surgical assistance tool, and for monitoring patient response to therapies.