Use of freeze-cracking in ontogenetic research in Macrostomum lignano (Macrostomida, Rhabditophora)
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A method for studying whole mount flatworm embryos based on freeze-cracking of the eggs is described. This method allows successful immunohistological and immunocytological studies of whole mount embryos. It does not require the use of sharpened needles or a microinjection system to puncture the eggshell. Moreover, this method is more practical and less time-consuming than classical puncturing and much cheaper than the use of a microinjection system. The advantages of this method are illustrated by results of several immunolocalisation experiments in the macrostomid flatworm Macrostomum lignano. The optimal procedure and crucial steps for this method are discussed.
KeywordsPlatyhelminthes Freeze-cracking Embryonic development Eggshell Immunohistochemistry
We are very grateful to Dr. Peter Ladurner (University of Innsbruck, Austria) for kindly providing the monoclonal antibodies. M.W. acknowledges the advice of Dr. Bernhard Egger and Dr. Volker Hartenstein regarding the stainings. This work was supported by IWT doctoral grants to M.W. and S.M. by the Institute for the Promotion of Innovation through Science and Technology in Flanders (IWT-Vlaanderen) and a FWO grant to M. B and K.D.M.
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