**2.4. Protein extraction by DNase/lithium chloride-dense sucrose homogenizationcoupled dichloromethane-methanol precipitation**

For each individual sample, a total of 500 mg placenta tissues were randomly chosen and pulverized in a mortar with pestle containing liquid nitrogen. Powdered placenta tissues were subjected to protein extraction using DNase/lithium chloride-dense sucrose, and then cleaned up with dichloromethane-methanol precipitation as described previously by Tan et al. [13].

These purified protein samples were stored at −80°C until 2D-GE was performed.
