als PDF - AGA

27. AGA-Kongress 2010 - Wien
P 5
Evaluation of cell viability in advanced cell-based therapies for cartilage repair
Wang W., Dono D., Duguay S.
Genzyme, Cambridge, United States
Purpose: Cell viability is fundamentally the most critical parameter for all cell-based therapeutic
products. However, viability testing remains a significant technical challenge for advanced 3-D cell
therapies and tissue engineering products. Here we report the development of a new, accurate and
reproducible assay to evaluate cell viability in matrix-induced autologous chondrocyte implantation
(MACI) for cartilage repair.
Method: The new method involves three simple steps:
1) the partial separation of cell-containing portion and non-cell-containing portion (conditioned media)
of the culture,
2) cell lysis and enzymatic reaction in the presence of fluorescent substrates, and
3) fluorescence measurement and a mathematic calculation of cell viability.
Results: The new method does not require cell recovery or control cells, and is not affected by donor
cell variability and cell distribution. It can be applied to both 2-D and 3-D cultures of human or animal
cells in all media types. The new assay demonstrates excellent accuracy (15%), precision (15%),
linearity (R 2 >0.95), and specificity (signal/noise ³ 3) over the full range of cell viabilities (0%-100%) and
a large range of cell densities (e.g. 8.75 e4 to 2.8 e6 per cm 2 membrane for MACI). The working cell
density range and medium type used in the assay can be easily adjusted using phenol red.
Conclusions: The new method successfully solved a long-standing technical challenge for advanced
cell therapies and tissue engineering products such as MACI. It is capable of measuring viability
accurately and precisely in all media types without interferences from the matrix material and donor
cell variability.
Keywords: Cell viability, cartilage, knee, cell therapy, tissue engineering
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