Fluorographic detection of tritiated glycopeptides and oligosaccharides separated on polyacrylamide gels: analysis of glycans from Dictyostelium discoideum glycoproteins |
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| Authors: | O P Das E J Henderson |
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| Affiliation: | 1. Advanced Polymer Materials Group, University Politehnica of Bucharest, SplaiulIndependenței, 313, 060042, Bucharest, Romania;2. Faculty of Medical Engineering, University Politehnica of Bucharest, Bucharest 011061, Romania;1. Department of Pharmacy, University of Salerno, Fisciano, SA, Italy;2. PhD Program in Drug Discovery and Development, University of Salerno (SA), Italy;3. Interdepartmental Research Center on Nuclear Magnetic Resonance for the Environment, Agro-food and New Materials (CERMANU), University of Naples Federico II, Portici, NA, Italy;4. Department of Agriculture, University of Naples Federico II, Portici, NA, Italy;1. BioX Centre, School of Biosciences and Bioengineering, Indian Institute of Technology Mandi, Himachal Pradesh 175075, India;2. School of Chemical Sciences and Advanced Materials Research Centre, Indian Institute of Technology Mandi, Himachal Pradesh 175075, India;3. University of Groningen, Zernike Institute for Advanced Materials, Polymer Science, Nijenborgh 4, 9747 AG Groningen, the Netherlands;4. The Silesian University of Science and Technology, Biotechnology Centre, Bolesława Krzywoustego 8, 44-100 Gliwice, Poland;5. School of Chemical Engineering, Yeungnam University, 280 Daehak-ro, Gyeongsan 38541, South Korea;6. University of Groningen, University Medical Center Groningen, Department of Biomedical Engineering, W. J. Kolff Institute, Antonius Deusinglaan 1, 9713 AV Groningen, the Netherlands;7. Division of Pediatric Orthopedic Surgery, Cincinnati Children''s Hospital Medical Center, Cincinnati, OH, USA;8. Department of Orthopedic Surgery, College of Medicine, University of Cincinnati, Cincinnati, OH, USA;9. Department of Biomedical Engineering, College of Engineering and Applied Sciences, University of Cincinnati, Cincinnati, OH, USA |
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| Abstract: | Previous workers have shown that oligosaccharides and glycopeptides can be separated by electrophoresis in buffers containing borate ions. However, normal fluorography of tritium-labeled structures cannot be performed because the glycans are soluble and can diffuse during equilibration with scintillants. This problem has been circumvented by equilibration of the gel with 2,5-diphenyloxazole (PPO) prior to electrophoresis. The presence of PPO in the gel during electrophoresis does not alter mobility of the glycopeptides and oligosaccharides. After electrophoresis, the gel is simply dried and fluorography performed. This allows sensitive and precise comparisons of labeled samples in parallel lanes of a slab gel and, since mobilities are highly reproducible, between different gels. The procedure is preparative in that after fluorography the gel bands can be quantitatively eluted for further study, without any apparent modification by the procedure. In this report, the procedure is illustrated by fractionation of both neutral and anionic glycopeptides produced by the cellular slime mold Dictyostelium discoideum. |
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