Mass Spectrometry Imaging of Organoids to Improve Preclinical Research.

Preclinical models are essential research tools before novel therapeutic or diagnostic methods can be applied to humans. These range from in vitro cell monocultures to vastly more complex animal models, but clinical translation to humans often fails to deliver significant results. Three-dimensional (3D) organoid systems are being increasingly studied to establish physiologically relevant in vitro platforms in a trade-off between the complexity of the research question and the complexity of practical experimental setups. The sensitivity and precision of analytical tools are yet another limiting factors in what can be investigated, and mass spectrometry (MS) is one of the most powerful analytical techniques available to the scientific community. Its innovative use to spatially resolve biological samples has opened many research avenues in the field of MS imaging (MSI). Here, this work aims to explore the current scientific landscape in the application of MSI on organoids, with an emphasis on their combined potential to facilitate and improve preclinical studies.

FluoMALDI Microscopy: Matrix Co-Crystallization Simultaneously Enhances Fluorescence and MALDI Imaging.

Here, the authors report that co-crystallization of fluorophores with matrix-assisted laser desorption/ionization (MALDI) imaging matrices significantly enhances fluorophore brightness up to 79-fold, enabling the amplification of innate tissue autofluorescence. This discovery facilitates FluoMALDI, the imaging of the same biological sample by both fluorescence microscopy and MALDI imaging. The approach combines the high spatial resolution and specific labeling capabilities of fluorescence microscopy with the inherently multiplexed, versatile imaging capabilities of MALDI imaging. This new paradigm simplifies registration by avoiding physical changes between fluorescence and MALDI imaging, allowing to image the exact same cells in tissues with both modalities. Matrix-fluorophore co-crystallization also facilitates applications with insufficient fluorescence brightness. The authors demonstrate  feasibility of FluoMALDI imaging with endogenous and exogenous fluorophores and autofluorescence-based FluoMALDI of brain and kidney tissue sections. FluoMALDI will advance structural-functional microscopic imaging in cell biology, biomedicine, and pathology.

RaMALDI: Enabling simultaneous Raman and MALDI imaging of the same tissue section.

Multimodal tissue imaging techniques that integrate two complementary modalities are powerful discovery tools for unraveling biological processes and identifying biomarkers of disease. Combining Raman spectroscopic imaging (RSI) and matrix-assisted laser-desorption/ionization (MALDI) mass spectrometry imaging (MSI) to obtain fused images with the advantages of both modalities has the potential of providing spatially resolved, sensitive, specific biomolecular information, but has so far involved two separate sample preparations, or even consecutive tissue sections for RSI and MALDI MSI, resulting in images with inherent disparities. We have developed RaMALDI, a streamlined, integrated, multimodal imaging workflow of RSI and MALDI MSI, performed on a single tissue section with one sample preparation protocol. We show that RaMALDI imaging of various tissues effectively integrates molecular information acquired from both RSI and MALDI MSI of the same sample, which will drive discoveries in cell biology, biomedicine, and pathology, and advance tissue diagnostics.

Prevalence and genetic characterization of Dirofilaria lutrae Orihle, 1965 in North American river otters (Lontra canadensis).

Dirofilaria lutrae Orihle, 1965 is a subcutaneous filarial worm found in North American river otters (Lontra canadensis). Previous studies reported the geographical range of this worm to include Louisiana, Florida, and North Carolina, USA. Although D. lutrae does not cause disease in otters, it must be distinguished from Dirofilaria immitis (Leidy, 1856), which causes heartworm disease in otters. The goal of this study was to determine the prevalence of D. lutrae in several states and to investigate the phylogenetic relationship of D. lutrae and other Dirofilaria species. Adult D. lutrae were collected from 32 of 40 (80%) otters from North Carolina (n = 38), South Carolina (n = 1), and Kentucky (n = 1), whereas all otters from Georgia (n = 1), Pennsylvania (n = 2), Arkansas (n = 184) and California (n = 1) were negative for D. lutrae. A portion of the cytochrome c oxidase I (COI) was amplified from nine North Carolina specimens and one South Carolina specimen. Sequence analysis indicated that all but one were identical to each other and D. lutrae was phylogenetically most closely related to D. immitis; however, few Dirofilaria sequences are available for comparison. Because other subcutaneous parasites may be detected in otters, parasites should be carefully examined to confirm their identity. Finally, given the high prevalence of D. lutrae in otters in some southeastern states, microfilaria found in blood samples of otters must be correctly identified because treatment for D. lutrae is generally not warranted and some pharmaceutical treatments can cause mortality in otters.