Macrocyclic Phage Display for Identification of Selective Protease Substrates.

Traditional methods for identifying selective protease substrates have primarily relied on synthetic libraries of linear peptides, which offer limited sequence and structural diversity. Here, we present an approach that leverages phage display technology to screen large libraries of chemically modified cyclic peptides, enabling the identification of highly selective substrates for a protease of interest. Our method uses a reactive chemical linker to cyclize peptides on the phage surface, while simultaneously incorporating an affinity tag and a fluorescent reporter.

Macrocyclic phage display for identification of selective protease substrates.

Traditional methods for identifying selective protease substrates have primarily relied on synthetic libraries of linear peptides, which offer limited sequence and structural diversity. Here, we present an approach that leverages phage display technology to screen large libraries of chemically modified cyclic peptides, enabling the identification of highly selective substrates for a protease of interest. Our method uses a reactive chemical linker to cyclize peptides on the phage surface, while simultaneously incorporating an affinity tag and a fluorescent reporter.

Recent advances in ratiometric fluorescence imaging of enzyme activity in vivo.

Among molecular imaging modalities that can monitor enzyme activity in vivo, optical imaging provides sensitive, molecular-level information at low-cost using safe and non-ionizing wavelengths of light. Yet, obtaining quantifiable optical signals in vivo poses significant challenges. Benchmarking using ratiometric signals can overcome dependence on dosing, illumination variability, and pharmacokinetics to provide quantitative in vivo optical data.

Shortwave infrared fluorescence imaging of peripheral organs in awake and freely moving mice.

Extracting biological information from awake and unrestrained mice is imperative to basic and pre-clinical research. Accordingly, imaging methods which preclude invasiveness, anesthesia, and/or physical restraint enable more physiologically relevant biological data extraction by eliminating these extrinsic confounders. In this article, we discuss the recent development of shortwave infrared (SWIR) fluorescent imaging to visualize peripheral organs in freely-behaving mice, as well as propose potential applications of this imaging modality in the neurosciences.

Protease Activated Probes for Real-Time Ratiometric Imaging of Solid Tumors.

Surgery is the preferred treatment option for most solid tumors. However, inaccurate detection of cancer borders leads to either incomplete removal of malignant cells or excess excision of healthy tissue. While fluorescent contrast agents and imaging systems improve tumor visualization, they can suffer from low signal-to-background and are prone to technical artifacts.

Shortwave infrared (SWIR) fluorescence imaging of peripheral organs in awake and freely moving mice.

Extracting biological information from awake and unrestrained mice is imperative to in vivo basic and pre-clinical research. Accordingly, imaging methods which preclude invasiveness, anesthesia, and/or physical restraint enable more physiologically relevant biological data extraction by eliminating these extrinsic confounders. In this article we discuss the recent development of shortwave infrared (SWIR) fluorescent imaging to visualize peripheral organs in freely-behaving mice, as well as propose potential applications of this imaging modality in the neurosciences.

Establishing design principles for emissive organic SWIR chromophores from energy gap laws.

Rational design of bright near and shortwave infrared (NIR: 700-1000 SWIR: 1000- 2000 nm) emitters remains an open question with applications spanning imaging and photonics. Combining experiment and theory, we derive an energy gap quantum yield master equation (EQME), describing the fundamental limits in SWIR quantum yields ( ) for organic chromophores. Evaluating the photophysics of 21 polymethine NIR/SWIR chromophores to parameterize the EQME, we explain the precipitous decline of past 900 nm through decreasing radiative rates and increasing nonradiative losses via high frequency vibrations relating to the energy gap.

Counterion Pairing Effects on a Flavylium Heptamethine Dye.

Polymethine fluorophores have facilitated the advance of biological and material sciences, due to their advantageous photophysical properties. However, the need to maintain a monomeric state can severely limit the utility and processability of dyes. High concentrations of fluorophore can lead to aggregation and negate the beneficial photophysical properties of monomers.

Photophysical Properties of Indocyanine Green in the Shortwave Infrared Region.

With the growing development of new contrast agents for optical imaging using near-infrared and shortwave infrared (SWIR) wavelengths, it is essential to have consistent bench-marks for emitters in these regions. Indocyanine green (ICG), a ubiquitous and FDA-approved organic dye and optical imaging agent, is commonly employed as a standard for photophysical properties and biological performance for imaging experiments at these wavelengths. Yet, its reported photophysical properties across organic and aqueous solvents vary greatly in the literature, which hinders its ability to be used as a consistent benchmark.

Bright Chromenylium Polymethine Dyes Enable Fast, Four-Color Imaging with Shortwave Infrared Detection.

Optical imaging within the shortwave infrared (SWIR, 1000-2000 nm) region of the electromagnetic spectrum has enabled high-resolution and high-contrast imaging in mice, non-invasively. Polymethine dyes, with their narrow absorption spectra and high absorption coefficients, are optimal probes for fast and multiplexed SWIR imaging. Here, we expand upon the multiplexing capabilities in SWIR imaging by obtaining brighter polymethine dyes with varied excitation wavelengths spaced throughout the near-infrared (700-1000 nm) region.

Shortwave infrared polymethine fluorophores matched to excitation lasers enable non-invasive, multicolour in vivo imaging in real time.

High-resolution, multiplexed experiments are a staple in cellular imaging. Analogous experiments in animals are challenging, however, due to substantial scattering and autofluorescence in tissue at visible (350-700 nm) and near-infrared (700-1,000 nm) wavelengths. Here, we enable real-time, non-invasive multicolour imaging experiments in animals through the design of optical contrast agents for the shortwave infrared (SWIR, 1,000-2,000 nm) region and complementary advances in imaging technologies.

Shortwave Infrared Imaging with J-Aggregates Stabilized in Hollow Mesoporous Silica Nanoparticles.

Tissue is translucent to shortwave infrared (SWIR) light, rendering optical imaging superior in this region. However, the widespread use of optical SWIR imaging has been limited, in part, by the lack of bright, biocompatible contrast agents that absorb and emit light above 1000 nm. J-Aggregation offers a means to transform stable, near-infrared (NIR) fluorophores into red-shifted SWIR contrast agents.

Flavylium Polymethine Fluorophores for Near- and Shortwave Infrared Imaging.

Bright fluorophores in the near-infrared and shortwave infrared (SWIR) regions of the electromagnetic spectrum are essential for optical imaging in vivo. In this work, we utilized a 7-dimethylamino flavylium heterocycle to construct a panel of novel red-shifted polymethine dyes, with emission wavelengths from 680 to 1045 nm. Photophysical characterization revealed that the 1- and 3-methine dyes display enhanced photostability and the 5- and 7-methine dyes exhibit exceptional brightness for their respective spectral regions.

Absence of radiographic abnormalities in nursemaid's elbows.

Background: Disagreement exists between physicians on the usefulness of a prereduction radiograph for diagnosis and treatment of nursemaid's elbows in children. Some evidence suggests that nursemaid's elbows have identifying features on radiographs. This study compares the radiographs of nursemaid's elbows to normal, control elbows in children and hypothesizes that differentiating features do not exist on radiograph.