Due to its intended application as a bone fixation and restoration technology, BoneFix is considered as an implantable medical device. As such, the BoneFix domains and their individual components must undergo careful biocompatibility tests to screen any potential toxicity on living cells and tissues, and thus ensure potential safe and effective use in humans.
At MINES Saint-Etienne, France, all the new chemical entities developed in the BoneFix consortium are being evaluated in vitro, following ISO standards, to ensure that they are harmless to cells. Dr Léa Dejob is leading this task and performs cell culture experiments and biochemical assays revealing any concentration dependent cytotoxicity or toxic by-products.
For instance, Live/Dead staining enables the direct discrimination of alive and dead cells via observation of the cultures under a fluorescent microscope. Briefly, after being exposed to BoneFix components, cells are incubated with a mixture of two dyes: one of them (calcein) can penetrate the cells, but will only be converted to a green-fluorescent compound by an enzyme found in living cells. The second fluorescent dye (ethidium homodimer III) can only penetrate cells that have lost their membrane integrity, and therefore labels dead cells in red by intercalating in their DNA. In parallel tests, cell death is also monitored by the quantification of lactate deshydrogenase (LDH) released by the cells in the culture media when they lose their membrane integrity. Another test is based on the ability of metabolically active cells to reduce a non-fluorescent dye (resazurin) into a highly fluorescent one (resorufin). Metabolic activity of the cells is then estimated by the intensity of fluorescence.
Once the cytocompatibility of the BoneFix building blocks is confirmed, preclinical proof of concept studies can be conducted by the other BoneFix consortium partners.
Comments