Abstract
The ligand of CD95, CD95L (also known as FasL or CD178), is a type II transmembrane protein that belongs to the Tumor Necrosis factor (TNF) family (Fig. 1a). This membrane-bound cytokine is mainly expressed at the surface of activated T lymphocytes and natural killer cells, where it is used as an apoptotic factor to eliminate infected and transformed cells (Strasser et al., Immunity 30:180–192, 2009).
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Notes
- 1.
MTT solution can be stored 6 months at 4 °C.
- 2.
Formation of precipitates can be followed by the observation of a cloudy solution as compared to a solution devoid of CaCl2, for example. Precipitates can be observed under a microscope.
- 3.
A concentration step can be performed either before or after ultracentrifugation, using centricon (Millipore) with a 10-kDa cut-off.
- 4.
Several gels can be transferred at the same time by placing them side-by-side on the semidry transfer platform.
- 5.
Red Ponceau can be used many times.
- 6.
Boyden chamber assay can be performed using either BT549 or MDA-MB 231 cell lines.
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Lepvrier, E., Martin, S., Collet, B. (2017). Production of the Non-apoptotic Metalloprotease-Cleaved CD95L and Its Cytotoxic Recombinant Counterpart Designed Ig-CD95L. In: Legembre, P. (eds) CD95. Methods in Molecular Biology, vol 1557. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6780-3_1
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DOI: https://doi.org/10.1007/978-1-4939-6780-3_1
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