Long-Term Detection of Fluorescently Labeled Human Mesenchymal Stem Cell In Vitro and In Vivo by Semi-Automated Microscopy

The use of seeded scaffolds in regenerative medicine is limited by the low survival of transplanted mesenchymal stem cells (MSC). Current approaches aim at improving cell viability but require an adequate long-term detection of the transplanted cells. Unfortunately, commonly performed labeling techn...

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Bibliographic Details
Published in:Tissue engineering. Part C, Methods Methods, 2012-02, Vol.18 (2), p.156-165
Main Authors: Polzer, Hans, Volkmer, Elias, Saller, Maximilian M., Prall, Wolf C., Haasters, Florian, Drosse, Inga, Anz, David, Mutschler, Wolf, Schieker, Matthias
Format: Article
Language:English
Subjects:
Algorithms
Animals
Automation
Bone Matrix - metabolism
Calcification, Physiologic
Cell Line
Cell Survival
Cell Tracking
Cellular biology
Flow Cytometry
Fluorescence
Fluorescent Dyes - metabolism
Green Fluorescent Proteins - metabolism
Humans
Mesenchymal Stromal Cells - cytology
Mesenchymal Stromal Cells - metabolism
Mice
Mice, Nude
Microscopy
Microscopy, Fluorescence - methods
Reproducibility of Results
Staining and Labeling
Stem cells
Tetrazolium Salts
Time Factors
Tissue engineering
Tissue Scaffolds - chemistry
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Summary:The use of seeded scaffolds in regenerative medicine is limited by the low survival of transplanted mesenchymal stem cells (MSC). Current approaches aim at improving cell viability but require an adequate long-term detection of the transplanted cells. Unfortunately, commonly performed labeling techniques have not been validated for this purpose, and studies often reveal inconclusive results. Consequently, we intended to identify the most suitable method for long-term detection of human MSC (hMSC) in vitro and in vivo. hMSC were labeled using the vital stainings PKH26 and carboxyfluorescein diacetate succinimidyl ester (CFDA-SE) as well as enhanced green fluorescent protein (eGFP) transduction. Metabolic activity and relative fluorescence intensity (RFI) were quantified in vitro over 21 days at 8 time points using standardized semi-automated microscopy and flow cytometry. In vivo , cell seeded scaffolds were subcutaneously implanted in nude mice, and RFI was analyzed over 42 days at 5 time points. In vitro , PKH26 and CFDA-SE significantly reduced metabolic activity. RFI of both stainings significantly decreased after 1 day and further faded to
ISSN:1937-3384
1937-3392
DOI:10.1089/ten.tec.2011.0275