Optimizing Human Synovial Fluid Preparation for Two-Dimensional Gel Electrophoresis |
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Authors: | Carl PC Chen Chih-Chin Hsu Wen-Lin Yeh Hsiu-Chu Lin Sen-Yung Hsieh Shih-Cherng Lin Tai-Tzung Chen Max JL Chen Simon FT Tang |
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Institution: | 1. Clinical & Molecular Genetics Unit, Institute of Child Health, University College London, 30 Guilford Street, London, WC1N 1EH, UK 2. Centre for Research in Biomedicine, Faculty of Health and Life Sciences, University of the West of England, Frenchay Campus, Coldharbour Lane, Bristol, BS16 1QY, UK 4. School of Biomedical and Biological Sciences, University of Plymouth, Drake Circus, Plymouth, PL4 8AA, UK 3. Fetal Medicine Unit, University College Hospital, London, NW1 2BU, UK
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Abstract: | Background Prenatal screening for Down Syndrome (DS) would benefit from an increased number of biomarkers to improve sensitivity and specificity. Improving sensitivity and specificity would decrease the need for potentially risky invasive diagnostic procedures. Results We have performed an in depth two-dimensional difference gel electrophoresis (2D DIGE) study to identify potential biomarkers. We have used maternal plasma samples obtained from first and second trimesters from mothers carrying DS affected fetuses compared with mothers carrying normal fetuses. Plasma samples were albumin/IgG depleted and expanded pH ranges of pH 4.5 - 5.5, pH 5.3 - 6.5 and pH 6 - 9 were used for two-dimensional gel electrophoresis (2DE). We found no differentially expressed proteins in the first trimester between the two groups. Significant up-regulation of ceruloplasmin, inter-alpha-trypsin inhibitor heavy chain H4, complement proteins C1s subcomponent, C4-A, C5, and C9 and kininogen 1 were detected in the second trimester in maternal plasma samples where a DS affected fetus was being carried. However, ceruloplasmin could not be confirmed as being consistently up-regulated in DS affected pregnancies by Western blotting. Conclusions Despite the in depth 2DE approach used in this study the results underline the deficiencies of gel-based proteomics for detection of plasma biomarkers. Gel-free approaches may be more productive to increase the number of plasma biomarkers for DS for non-invasive prenatal screening and diagnosis. |
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