Double labelling of human umbilical cord mesenchymal stem cells with Gd‐DTPA and PKH26 and the influence on biological characteristics of hUCMSCs

The aim of this study was to determine whether double labelling of human umbilical cord mesenchymal stem cells (hUCMSCs) with gadolinium‐diethylene triamine penta‐acetic acid (Gd‐DTPA) and PKH26 influences their biological characteristics. A tissue adherence technique was used to separate and purify the hUCMSCs and flow cytometry was performed to detect the surface markers expressed on them. Gd‐DTPA and PKH26 were used to label the stem cells and MRI and fluorescence microscopy were used to detect the double‐labelled hUCMSCs. A MTT assay was used to delineate the growth curve. Transmission electron microscopy (TEM) and atomic force microscopy were used to demonstrate the ultrastructural features of the hUCMSCs. Flow cytometry showed that hUCMSCs highly expressed CD29, CD90, CD44 and CD105. No expression of CD31, CD34 and CD45 was detected. Very low expression of HLA‐DR and CD40 was detected. Atomic force microscopy showed these cells were long, spindle shaped, and the cytoplasm and nucleus had clear boundaries. After double labelling, TEM showed Gd particles aggregated in the cytoplasm in a cluster pattern. The proliferation activity, cell cycle, apoptosis and differentiation of the stem cells were not influenced by double labelling. Thus a tissue adherence technique is helpful to separate and purify hUCMSCs effectively; and Gd‐DTPA and PKH26 are promising tracers in the investigation of migration and distribution of hUCMSCs in vivo.