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Figure 1: Flow cytometry of HEK 293T cells transfected with FLAG tagged mutated mouse caspase-8.Method: MAbs 1G12 and 3B10 both detect overexpressed FLAG-epitope-tagged pro-caspase-8 (active-site-mutant) by flow cytometry in permeabilized cells. As a positive control staining with a MAb to FLAG was performed. As a negative control an isotype control was employed.
Figure 2: MAbs 1G12 and 3B10 both detect pro-caspase-8 in MEFs from WT mice, but not in MEFs from caspase-8-/- mice. Several smaller bands detected in the caspase-8-/- MEFs, correspond to truncated forms of caspase-8 made in the caspase-8-/- mice since only exons 1 and 2 of mouse caspase-8 were deleted in these knock-out mice and not the region encoding the p18 subunit.Note: Extra bands marked by * are only seen in lysates from caspase-8-/- MEFs and not in lysates from any WT cell lines or mouse WT tissue. Both MAbs to caspase-8 (mouse) do not recognize human caspase-8, whereas endogenous caspase-8 can be efficiently detected in various mouse cell lines. Upon an apoptotic stimulus e.g. by cross-linked rhsFasL both MAbs to caspase-8 (mouse) do also recognize the cleaved active p18 subunit of mouse caspase-8 in addition to the caspase-8 precursor.
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RUO – Research Use Only |
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