Answer: (B) Differential centrifugation

Differential centrifugation is the standard first step to separate water-soluble proteins in the supernatant from insoluble cellular debris like nuclei and unbroken cells after cell lysis.

Option Breakdown

Electroporation (A)

Electroporation uses electric pulses to create temporary pores in cell membranes for DNA/protein delivery.​
It introduces molecules into intact cells but does not separate soluble from insoluble protein fractions.​

Differential Centrifugation (B)

Low-speed centrifugation (500-20,000g) pellets heavy insoluble material (nuclei, mitochondria) while soluble cytosolic proteins remain in supernatant.
This stepwise process by size/density yields clarified lysate for downstream purification.​

Density-Gradient Centrifugation (C)

Sucrose or Percoll gradients separate organelles by buoyant density after initial differential steps.
It’s a refinement technique, not the initial separation of soluble vs. insoluble fractions.​

Western Blotting (D)

Western blotting detects specific proteins via antibody binding after gel electrophoresis.​
This analytical method follows purification; it cannot perform physical separation.​

The most common initial step to separate water soluble proteins insoluble material uses differential centrifugation on homogenized cells.

How Differential Centrifugation Works

Cells lyse via homogenization, then spin at 800g to pellet unbroken cells/nuclei; supernatant spins at 10,000g for mitochondria.
Soluble proteins stay supernatant-ready for chromatography or dialysis.​

Why Not Other Methods?

Electroporation disrupts membranes for uptake, not fractionation.​
Density gradients polish fractions post-differential; Western analyzes purified samples.

Lab Protocol Essentials

Start 300-1000g (5-10 min, 4°C) for debris; progress to higher speeds.
Supernatant contains ~80% cytosolic water-soluble proteins.​

This establishes B as the routine first purification step.​