Detecting seasonal variation of antifreeze protein distribution in Rhagium mordax using immunofluorescence and high resolution microscopy |
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| Institution: | 1. Laboratory of Pharmaceutical Biotechnology, Ghent University, 9000 Ghent, Belgium;2. CNRS, IPBS (Institut de Pharmacologie et Biologie Structurale), Toulouse, France;3. Université de Toulouse, UPS, IPBS, Toulouse, France;4. Laboratoire des Sciences du Numérique de Nantes, Nantes University, 44035 Nantes, France;5. Research Institute of Molecular Pathology (IMP), Vienna Biocenter (VBC), Campus-Vienna-Biocenter 1, 1030 Vienna, Austria;6. IMBA Institute of Molecular Biotechnology of the Austrian Academy of Sciences, Vienna Biocenter (VBC), Dr. Bohr-Gasse 3, 1030 Vienna, Austria;7. Department of Biochemistry and Molecular Biology, VILLUM Center for Bioanalytical Sciences, University of Southern Denmark, Campusvej 55, 5230 Odense, Denmark;8. Medizinisches Proteom-Center, Ruhr-University Bochum, Universitätsstr. 150, 44801 Bochum, Germany;9. KG Jebsen Center for Diabetes Research, Department of Clinical Science, University of Bergen, Norway;10. Center for Medical Genetics and Molecular Medicine, Haukeland University Hospital, Bergen, Norway;11. Bioinformatics Research Group, University of Applied Sciences Upper Austria, Softwarepark 11, 4232 Hagenberg, Austria |
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| Abstract: | Larvae of the blackspotted pliers support beetle, Rhagium mordax, were collected monthly, for the duration of 2012 and fixed. The larvae were embedded in paraffin wax and sectioned. Using fluorophore-coupled antibodies specific to the R. mordax antifreeze protein, RmAFP1, sections were visualised with UV reflected light microscopy. An automated software analysis method was developed in order to discard autofluorescence, and quantify fluorescence from bound antibodies. The results show that R. mordax cuticle and gut exhibit a higher degree of fluorophore-bound fluorescence during summer, than in the cold months. It is hypothesised that R. mordax stores RmAFP1 in, or near, the fat body during times when freeze avoidance is not needed. |
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| Keywords: | Antifreeze protein Immunofluorescence Microscopy Detection Cross section Image analysis DAPI"} {"#name":"keyword" "$":{"id":"kwrd0015a"} "$$":[{"#name":"text" "_":"4′ 6-diamidino-2-phenylindole (blue 461 nm) FITC"} {"#name":"keyword" "$":{"id":"kwrd0025a"} "$$":[{"#name":"text" "_":"Fluorescein isothiocyanate (green 518 nm) TRITC"} {"#name":"keyword" "$":{"id":"kwrd0035a"} "$$":[{"#name":"text" "_":"Rhodamine (red 580 nm) |
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