Cryo-ultramicrotomy and Mass Spectrometry Imaging Analysis of Nudibranch Microstructures

Hamilton, Brett; Chan, Weili; Cheney, Karen L.; Sullivan, Rodney N.; Floetenmeyer, Matthias; Garson, Mary J.; Wepf, Roger

doi:10.1021/jasms.1c00254

Cited by 7 publications

(6 citation statements)

References 16 publications

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“…Spatial omics studies are rare in marine invertebrates (Geier et al, 2020; Goto-Inoue et al, 2020; Hamilton et al, 2022), and the difficulty in obtaining intact tissue sections in the organism’s natural shape is a major hurdle. Through the combination of anesthetizing, embedding and cryofilm application, the cryosectioning workflow presented produced intact tissue sections for a soft-bodied (sea anemone E. diaphana ) and a hard-bodied (coral G. fascicularis ) cnidarian in their natural shape (Fig.…”

Section: Resultsmentioning

confidence: 99%

Spatial metabolomics for symbiotic marine invertebrates

Chan

Rudd

Oppen

2022

Preprint

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Microbial symbionts frequently localize within specific body structures or cell types of their multicellular hosts. This spatiotemporal niche is critical to host health, nutrient exchange and fitness. Measuring host-microbe metabolite exchange has conventionally relied on tissue homogenates, eliminating dimensionality and dampening analytical sensitivity. We have developed a mass spectrometry imaging (MSI) workflow for a soft- and hard-bodied cnidarian animal capable of revealing the host and symbiont metabolome in situ, without the need for a priori isotopic labelling or skeleton decalcification. The MSI method provides critical functional insights that cannot be gleaned from bulk tissue analyses or other presently available spatial methods. We show that cnidarian hosts may regulate microalgal symbionts acquisition and rejection through specific ceramides distributed throughout the tissue lining the gastrovascular cavity; once resident, symbionts reside in light-exposed tentacles to generate photosynthate. These spatial patterns reveal how symbiont identity can drive host metabolism.

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Section: Resultsmentioning

confidence: 99%

Spatial metabolomics for symbiotic marine invertebrates

Chan

Rudd

Oppen

2022

Preprint

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“…Although MSI is well integrated into medical studies and other biological research such as on plants (Gupta et al, 2019;Nakabayashi et al, 2019) and invertebrates (Kopp et al, 2015b;Yang et al, 2020;Bourceau et al, 2022;Hamilton et al, 2022), only a few non-medical studies have explored its application in hostsymbiont interactions. The ability to study hundreds to thousands of metabolites in situ via MSI makes this a powerful tool for researchers to decipher host-microbe interactions.…”

Section: Introductionmentioning

confidence: 99%

Spatial metabolomics for symbiotic marine invertebrates

2023

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Microbial symbionts frequently localize within specific body structures or cell types of their multicellular hosts. This spatiotemporal niche is critical to host health, nutrient exchange, and fitness. Measuring host–microbe metabolite exchange has conventionally relied on tissue homogenates, eliminating dimensionality and dampening analytical sensitivity. We have developed a mass spectrometry imaging workflow for a soft- and hard-bodied cnidarian animal capable of revealing the host and symbiont metabolome in situ, without the need for a priori isotopic labelling or skeleton decalcification. The mass spectrometry imaging method provides critical functional insights that cannot be gleaned from bulk tissue analyses or other presently available spatial methods. We show that cnidarian hosts may regulate microalgal symbiont acquisition and rejection through specific ceramides distributed throughout the tissue lining the gastrovascular cavity. The distribution pattern of betaine lipids showed that once resident, symbionts primarily reside in light-exposed tentacles to generate photosynthate. Spatial patterns of these metabolites also revealed that symbiont identity can drive host metabolism.

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“…Chen et al used a sucrose solution as an embedding medium, and used a cryoultramicrotome to microtome the polymer film into ultrathin sections of 50 nm thickness for the transmission electron microscopy characterization [ 28 ]. The sample had to be trimmed into blocks of a few hundred microns or even smaller to fit the sectioning range of the cryoultramicrotome [ 29 , 30 , 31 ]. However, the resultant cross-sectional area was too small for the SEM analysis.…”

Section: Resultsmentioning

confidence: 99%

Preparation of Cross-Sectional Membrane Samples for Scanning Electron Microscopy Characterizations Using a New Frozen Section Technique

Ren

Zhang

et al. 2023

Membranes

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Characterization of the cross-sectional morphologies of polymeric membranes are critical in understanding the relationship of structure and membrane separation performances. However, preparation of cross-sectional samples with flat surfaces for scanning electron microscopy (SEM) characterizations is challenging due to the toughness of the non-woven fabric support. In this work, a new frozen section technique was developed to prepare the cross-sectional membrane samples. A special mold was self-designed to embed membranes orientationally. The frozen section parameters, including the embedding medium, cryostat working temperature, and sectioning thickness were optimized. The SEM characterizations demonstrated that the frozen section technique, using ultrapure water as the embedding medium at a working temperature of −30 °C and a sectioning thickness of 0.5 µm, was efficient for the preparation of the membrane samples. Three methods of preparation for the cross-sectional polymeric membranes, including the conventional liquid nitrogen cryogenic fracture, the broad ion beam (BIB) polishing, and the frozen section technique were compared, which showed that the modified frozen section method was efficient and low cost. This developed method could not only accelerate the development of membrane technology but also has great potential for applications in preparation of other solid samples.

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Cryo-ultramicrotomy and Mass Spectrometry Imaging Analysis of Nudibranch Microstructures

Cited by 7 publications

References 16 publications

Spatial metabolomics for symbiotic marine invertebrates

Spatial metabolomics for symbiotic marine invertebrates

Spatial metabolomics for symbiotic marine invertebrates

Preparation of Cross-Sectional Membrane Samples for Scanning Electron Microscopy Characterizations Using a New Frozen Section Technique

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