Development of novel monoclonal antibodies that de

Authors: Ditte C Andersen Angela Kortesidis Andrew C W Zannettino Irina Kratchmarova Li Chen Ole N Jensen Børge Teisner Stan Gronthos Charlotte H Jensen Moustapha Kassem

Publish Date: 2011/05/23

Volume: 32, Issue: 2, Pages: 133-142

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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 835 11 and 85 of primary cultures of hMSC were double positive for STRO1 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 massspectrometric analysis we identified the antigen recognised by DJ3 as CD44 whereas DJ9 and DJ18 recognized HLADRB1 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 STRO1+/−/Collagen VI− sorted hMSC contained fewer differentiated alkaline phosphatase+ cells compared to STRO1+/−/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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