Western Blot Analysis of Parasite Proteins

Parasite proteins isolated from different subcellular fractionations were separated on SDS-polyacrylamide gels and were transferred to polyvinylidene difluoride (PVDF) membranes as previously described (48 (link)). The primary antibodies used were rabbit anti-PfRad51 (1:5,000 dilution) (25 (link)), rabbit anti-PfBlm (1:5,000) (27 (link)), rabbit anti-PfalMre11 (1:5,000) (26 (link)), rabbit anti-HsHistone 3 (1:5,000; Imperial Life Sciences), mouse anti-cytochrome c (1:5,000; Abcam), rabbit anti-GFP (1:5,000; Abcam), and mouse anti-GAPDH (1:5,000; Abcam). The membranes were washed with Tris-buffered saline with Tween 20 (TBS-T) and treated with horseradish peroxidase (HRP)-conjugated anti-mouse (1:10,000; Santa Cruz Biotechnology Inc.) and anti-rabbit (1:10,000; Promega) secondary antibody for 2 h at 4°C. After washes with TBS-T, membranes were developed with a chemiluminescent horseradish peroxidase (HRP) substrate (SuperSignal West Pico Plus; Thermo Scientific) and imaged by using a ChemiDoc Touch imaging system from Bio-Rad.