Abstract
BackgroundGelatinous zooplankton can be difficult to preserve morphologically due to unique physical properties of their cellular and acellular components. The relatively large volume of mesoglea leads to distortion of the delicate morphology and poor sample integrity in specimens prepared with standard aldehyde or alcohol fixation techniques. Similar challenges have made it difficult to extend standard laboratory methods such as in situ hybridization to larger juvenile ctenophores, hampering studies of late development.ResultsWe have found that a household water repellant glass treatment product commonly used in laboratories, Rain-X®, alone or in combination with standard aldehyde fixatives, greatly improves morphological preservation of such delicate samples. We present detailed methods for preservation of ctenophores of diverse sizes compatible with long-term storage or detection and localization of target molecules such as with immunohistochemistry and in situ hybridization and show that this fixation might be broadly useful for preservation of other delicate marine specimens.ConclusionThis new method will enable superior preservation of morphology in gelatinous specimens for a variety of downstream goals. Extending this method may improve the morphological fidelity and durability of museum and laboratory specimens for other delicate sample types.
Highlights
Gelatinous zooplankton can be difficult to preserve morphologically due to unique physical properties of their cellular and acellular components
Rain-X® fixation of cydippid and lobate stage Mnemiopsis Samples fixed with Rain-X® are consistently preserved at a range of body sizes (Fig. 1) as opposed to more standard methods such as aldehyde fixation in which a majority of samples may be shriveled or destroyed (Supplemental Figure 1 A-C)
This is especially valuable for precious specimens and serves to streamline protocols such as in situ hybridization and immunofluorescence staining since more interpretable, publication-quality images can be obtained from the same sample size
Summary
Gelatinous zooplankton can be difficult to preserve morphologically due to unique physical properties of their cellular and acellular components. The contrasting physical properties of the mesoglea and fragile tissue layers is a long-standing, well known problem to laboratory biologists, field ecologists, and museum curators interested in the long and short term preservation of these kinds of animals [1] This problem has limited developmental biologists and others interested in using downstream molecular techniques to study various aspects of these animals’ biology. We serendipitously have developed a new protocol to fix whole animals that better preserves overall morphology than existing methods and is compatible with a broad range of powerful molecular techniques These include immunohistochemistry, small molecule labeling (such as with EdU), and in situ hybridization, techniques that are often difficult to interpret with morphological degradation. We hope this technique will be successful in other difficultto-fix jellies
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