Quantifying Protein Expression by Western Blot

The expression of marker proteins was tested using western blotting as previously described [24 (link)]. Cellular lysates were prepared 30 min and 24 h post-IR according to standard procedures. Forty μg of protein per lane were separated using 4–12% SDS-polyacrylamide pre-cast gels (Invitrogen, Karlsruhe, Germany) and transferred to nitrocellulose membranes according to the manufacturer’s prescriptions. The levels of protein expression were quantified using Image J (Wayne Rasband, National Institutes of Health, Bethesda, MD) and normalized to β-actin intensity. Experiments were repeated at least three times, unless otherwise noted.