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By Kate Monica

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British Soap AwardsThe moment pregnant Jacqueline Jossa's smile drops as she's forced to walk the British Soap Awards red carpet aloneThe heavily pregnant actress is currently separated from her husband Dan Osborne as she prepares to give birth
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Western blotting

Heart and cell homogenates (30 ?g protein), or isolated cytosolic (20 ?g protein), mitochondrial (15 ?g protein), and nuclear (20 ?g protein) fractions were electrophoresed on a 4–15% Mini-Protein TGX gel using a Mini-PROTEAN electrophoresis chamber (Bio-Rad), and proteins were transferred to a PVDF membrane using a Criterion Trans-blot System (Bio-Rad). Membranes were blocked with 5% non-fat dry milk (NFDM) in 50 mM Tris-HCl (pH 7.5) / 150 mM NaCl (TBS), washed with TBS-0.1% Tween 20 (TBST) and TBS, and incubated overnight at 4°C with antibody clones listed in Table C of S1 Table. Antibodies from Santa Cruz were used at 1:200 dilutions, and all other antibodies were used at 1:1000 dilution in TBST-5% NFDM. Membranes were washed with TBST and TBS, incubated with HRP-conjugated secondary antibody (1: 10,000 dilution, Southern Biotech, Birmingham AL), and images were acquired by using an Image Quant LAS4000 system (GE Healthcare, Pittsburgh MA). Immunoblots were subjected to Ponceau S staining to confirm equal loading and transferring of samples. Densitometry analysis of protein bands was performed using a Fluorchem HD2 Imaging System (Alpha Innotech, San Jose CA), and normalized against GAPDH (tissue homogenates and cytosolic fractions), COIV (mitochondrial fractions) or Lamin B (nuclear fractions).