Hyperspectral Multiplexed Biological Imaging of Nanoprobes Emitting in the Short-Wave Infrared Region

Abstract

Optical bioimaging with exogenous luminophores emitting in short-wave infrared spectral region (SWIR, ~ 1000–1700 nm) is a rapidly developing field, and the development of multiple SWIR-photoluminescent nanoprobes has recently been reported. In this regard, hyperspectral imaging (HSI), combined with unmixing algorithms, is a promising tool that can allow for efficient multiplexing of the SWIR-emitting nanoagents by their photoluminescence (PL) spectral profiles. The SWIR HSI technique reported here is developed to multiplex two types of nanoprobes: polymeric nanoparticles doped with organic dye (PNPs) and rare-earth doped fluoride nanoparticles (RENPs). Both types of nanoprobes exhibit PL in the same spectral range (~ 900–1200 nm), which hinders spectral separation of PL with optical filters and limits possibilities for their multiplexed imaging in biological tissues. By applying SWIR HSI, we exploited differences in the PL spectral profiles and achieved the spectrally selective and sensitive imaging of the PL signal from every type of nanoparticles. Unmixing of acquired data allowed for multiplexing of the spectrally overlapping nanoprobes by their PL profile. Both quantitative and spatial distribution for every type of nanoparticles were obtained from their mixed suspensions. Finally, the SWIR HSI technique with unmixing protocol was applied to in vivo imaging of mice subcutaneously injected with PNPs and RENPs. The applicability of hyperspectral techniques to multiplex nanoprobes in the in vivo imaging was successfully demonstrated.

Core–shell polymeric nanoparticles co-loaded with photosensitizer and organic dye for photodynamic therapy guided by fluorescence imaging in near and short-wave infrared spectral regions

“…Though JB7-08 and JB17-08 are virtually non-fluorescent in aqueous solutions, their fluorescence is restored through post-loading to the polymeric matrix of PS-PNIPAM nanoparticles [24]. As a result, the dye-loaded nanoparticles exhibit fluorescence in ~ 900-1100 nm spectral range under excitation at ~ 800 nm [31]. These spectral characteristics suggest that JB 7-08 and JB17-08 dyes are promising candidates as imaging agents co-loaded with HPPH in PAA-based NPs.…”

Phytoglycogen Encapsulation of Lanthanide‐Based Nanoparticles as an Optical Imaging Platform with Therapeutic Potential

“…[ 53 ] This technique is particularly attractive in combination with optical multiplexing as it can significantly increase the throughput of (bio)imaging by tracking multiple targets, such as fluorescent proteins or luminescent nanoparticles. [ 54,55 ] The bright field optical microscopy image (Figure 2A2) shows a region containing UCNP‐PGLY beads of similar morphologies. However, when hyperspectral mapping was performed at the regions of interest indicated in green, blue and cyan (magnified in Figure 2B/C1–4 for better visualization), the characteristic spectral profiles of UCNP‐2, UCNP‐3, and UCNP‐4 were unequivocally identified (Figure 2D1–4).…”

Non-invasive synchronous monitoring of neutrophil migration using whole body near-infrared fluorescence-based imaging

“…To overcome these challenges, we have optimised an in vivo dual fluorescent imaging method. There are literature reports which demonstrate in vivo multiplexing, the experiments have been performed in BALB/c or nude mice to avoid fine black fur and skin pigmentation 12,29,30 . Melanin skin pigmentation significantly attenuates signal detection 18 , hypothesised to be due to the high absorption coefficient associated with melanin 31 .…”