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An Image-Based Assay of Endothelial Cell Tube Formation as a Model of Angiogenesis

he development of specific assays. The software allows cellular debris or non-connected tube fragments to be removed prior to data analysis. The data generated is contained in text files and easily analyzed using BD Image Data Explorer (Cat. No. 341039).

Control images ( Figure 3 , left panel) obtained with the BD Pathway Bioimager show distinct development of tubes indicating that endothelial cells indeed do resemble blood vessels. The image in Figure 1 was acquired using a 10 objective and shows very clear development of endothelial cell-derived tubules. These tubules mostly originate at aggregates of cells and appear to actively connect with other tubules or aggregates. The total number of tubes formed ( Figure 3 and Figure 4 ) is an important measurement parameter since the underlying signaling and gene expression processes which mediates tube formation may offer an attractive leverage point for drug development.

Discussion

The advent of automated imaging platforms such as the BD Pathway Bioimager allow the migration of microscopebased assays into the drug discovery and systems biology arena. For this to succeed, images not only need to be acquired in a highly reproducible and hands-off manner but must also be analyzed with minimal user intervention. We present here such an assay consisting of human endothelial cells, 96-well coated plates, an automated imaging system and data analysis software. Confocal microscopy combined with the collapsed stack feature of the BD Pathway Bioimager enables analysis of angiogenic tubules in a 3 dimensional matrix that better resembles the native physiological environment. Alternative methods using non-confocal systems are being used where the matrix is dried out overnight to collapse the cells into a flat plane this can not be done on live cells and may create significant distortions in the fragile branching structure of the samp
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