Abstract
.The observed behavior of short-wave infrared (SWIR) light in tissue, characterized by relatively low scatter and subdiffuse photon transport, has generated considerable interest for the potential of SWIR imaging to produce high-resolution, subsurface images of fluorescence activity in vivo. These properties have important implications for fluorescence-guided surgery and preclinical biomedical research. Until recently, translational efforts have been impeded by the conventional understanding that fluorescence molecular imaging in the SWIR regime requires custom molecular probes that do not yet have proven safety profiles in humans. However, recent studies have shown that two readily available near-infrared (NIR-I) fluorophores produce measurable SWIR fluorescence, implying that other conventional fluorophores produce detectable fluorescence in the SWIR window. Using SWIR spectroscopy and wide-field SWIR imaging with tissue-simulating phantoms, we characterize and compare the SWIR emission properties of eight commercially available red/NIR-I fluorophores commonly used in preclinical and clinical research, in addition to a SWIR-specific fluorophore. All fluorophores produce measurable fluorescence emission in the SWIR, including shorter wavelength dyes such as Alexa Fluor 633 and methylene blue. This study is the first to report SWIR fluorescence from six of the eight conventional fluorophores and establishes an important comparative reference for developing and evaluating SWIR imaging strategies for biomedical applications.
Highlights
Reports of fluorescence imaging in the short-wave infrared (SWIR) regime,[1,2,3] known as the near-infrared II (NIR-II) window, have generated substantial interest in the biomedical optics community
With the exception of indocyanine green (ICG), which was dissolved in deionized water, all fluorophores were diluted in phosphate-buffered saline (PBS)
Fluorophores with absorbance and emission peaks at shorter wavelengths—AF 633, AF 647, methylene blue, IRDye 680RD, and IRDye 700DX—generated the lowest signal intensities in the SWIR window, and the highest fluorescence intensities were observed for the longer wavelength fluorophores
Summary
Reports of fluorescence imaging in the short-wave infrared (SWIR) regime,[1,2,3] known as the near-infrared II (NIR-II) window, have generated substantial interest in the biomedical optics community. A more significant barrier arose from the conventional assumption that specialized molecular constructs were required for SWIR fluorescence imaging These molecular probes can be difficult to obtain, usually manufactured in individual research labs (with a few commercial exceptions), and, importantly, do not yet have pharmacokinetic and safety profiles necessary for mid-to-late-stage translational research. These barriers have generally limited SWIR imaging to animal research in labs with appropriate expertise
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