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Development of novel monoclonal antibodies that define differentiation stages of human stromal (mesenchymal) stem cells

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Molecules and Cells

Abstract

Human mesenchymal stem cells (hMSC) are currently being introduced for cell therapy, yet, antibodies specific for native and differentiated MSCs are required for their identification prior to clinical use. Herein, high quality antibodies against MSC surface proteins were developed by immunizing mice with hMSC, and by using a panel of subsequent screening methods. Flow cytometry analysis revealed that 83.5, 1.1, and 8.5% of primary cultures of hMSC were double positive for STRO-1 and either of DJ 3, 9, and 18, respectively. However, none of the three DJ antibodies allowed enrichment of clonogenic hMSC from BMMNCs as single reagents. Using mass-spectrometric analysis, we identified the antigen recognised by DJ3 as CD44, whereas DJ9 and DJ18 recognized HLA-DRB1 and Collagen VI, respectively. The identified proteins were highly expressed throughout in vitro osteogenic- and adipogenic differentiation. Interestingly, undifferentiated cells revealed a sole cytoplasmic distribution pattern of Collagen VI, which however changed to an extracellular matrix appearance upon osteogenic- and adipogenic differentiation. In relation to this, we found that STRO-1+/−/Collagen VI− sorted hMSC contained fewer differentiated alkaline phosphatase+ cells compared to STRO-1+/−/Collagen VI+ hMSC, suggesting that Collagen VI on the cell membrane exclusively defines differentiated MSCs. In conclusion, we have generated a panel of high quality antibodies to be used for characterization of MSCs, and in addition our results may suggest that the DJ18 generated antibody against Collagen VI can be used for negative selection of cultured undifferentiated MSCs.

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Author notes

  1. Ditte C. Andersen & Charlotte H. Jensen

    Present address: Laboratory of Molecular and Cellular Cardiology, Department of Clinical Biochemistry and Pharmacology, Odense University Hospital, Odense, Denmark

Authors and Affiliations

  1. Department of Cancer and Inflammation Research, University of Southern Denmark, Odense, Denmark

    Ditte C. Andersen, Børge Teisner & Charlotte H. Jensen

  2. Mesenchymal Stem Cell Group, Department of Haematology, Institute of Medical and Veterinary Science/Hanson Institute, Adelaide, South Australia, Australia

    Angela Kortesidis & Stan Gronthos

  3. Myeloma Research Laboratory, Department of Haematology, Institute of Medical and Veterinary Science/Hanson Institute, Adelaide, South Australia, Australia

    Andrew C. W. Zannettino

  4. Center for Experimental BioInformatics, Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, Denmark

    Irina Kratchmarova & Ole N. Jensen

  5. Department of Endocrinology and Metabolism, Odense University Hospital, Odense, Denmark

    Li Chen & Moustapha Kassem

  6. Stem Cell Unit, Department of Anatomy, College of Medicine, King Saud Univeristy, Riyadh, KSA

    Moustapha Kassem

Authors
  1. Ditte C. Andersen
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  2. Angela Kortesidis
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Correspondence to Ditte C. Andersen.

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Andersen, D.C., Kortesidis, A., Zannettino, A.C.W. et al. Development of novel monoclonal antibodies that define differentiation stages of human stromal (mesenchymal) stem cells. Mol Cells 32, 133–142 (2011). https://doi.org/10.1007/s10059-011-2277-7

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  • DOI: https://doi.org/10.1007/s10059-011-2277-7

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