Labeling Mesenchymal Stromal Cells with PKH26 or VybrantDilSignificantly Diminishes their Migration, but does not affect theirViability, Attachment, Proliferation and Differentiation Capacities

Fluorescent dyes such as PKH26 and VybrantDil facilitate rapid and simple labeling of cells for later detection in various assays. But covering the cell surface with lipophilic fluorescent dyes may impair cellular functions. We therefore investigated the effects of PKH26 and VybrantDil on viability, proliferation, differentiation, attachment and migration of human mesenchymal stromal cells (MSCs) in vitro. To this end, MSCs were harvested from bone marrow of 12 individuals, expanded employing methods compliant to good manufacturing practice, and stained with PKH26 or VybrantDil. MSCs without label served as controls. The intensity of fluorescent staining as function of label and incubation time was investigated. Viability and proliferation were enumerated by cell counting. The osteogenic and adipogenic differentiations of MSCs were explored by cytochemical staining and transcript analyses, the cell migration and attachment by specific in vitro assays. We report that labeling of human MSCs with PKH26 yielded brighter signals facilitating prolonged detection compared to VybrantDil. No significant effects of PKH26 and VybrantDil were recorded when the viability, proliferation, attachment, osteogenic and adipogenic differentiation of human MSCs were investigated. But loading cells with PKH26 or VybrantDil significantly diminished the migration of the MSCs in vitro. We conclude that analyses depending on cellular migration may be biased when the cells are loaded with these lipophilic dyes.