Centrifugation and filtration are the most common methods for separating solids from liquids in the laboratory. They are used across all disciplines — from pelleting bacteria to clarifying protein extracts to filtering HPLC mobile phases.

Centrifugation Principles

Centrifugation uses centrifugal force to sediment particles based on their size, shape, and density. The relative centrifugal force (RCF), expressed in × g, is calculated as:

RCF = 1.118 × r × (RPM/1000)²

where r is the rotor radius in millimeters. Always use RCF (× g) rather than RPM when comparing protocols between different centrifuges, since the radius varies by rotor.

Rotor Types

• Fixed-angle rotors: tubes are held at a fixed angle (typically 25–45°). Pellets form on the side of the tube. Good for most pelleting applications.
• Swinging-bucket rotors: tubes hang vertically during loading and swing out horizontally during centrifugation. Pellets form at the tube bottom, giving better separation for density gradients.
• Vertical rotors: tubes remain vertical throughout. Used for rapid separations in density gradient ultracentrifugation.
• Microcentrifuges: small fixed-angle rotors for 1.5–2 mL tubes, common for DNA/RNA work.

Choosing Speed and Time

The sedimentation rate depends on particle size — larger particles pellet at lower speeds. Typical protocols:

• Low-speed (500–2000 × g): pelleting cells, large debris.
• Medium-speed (5000–15000 × g): pelleting organelles, bacteria, protein precipitates.
• High-speed (20000–100000 × g): pelleting microsomes, small vesicles.
• Ultracentrifugation (>100000 × g): pelleting viruses, ribosomes, macromolecular complexes.

Balancing and Safety

Always balance tubes by mass (not volume) to within 0.1 g for high-speed rotors and 0.5 g for low-speed rotors. Load opposing tubes with identical tube types and fill volumes. Never exceed the maximum speed rating for a rotor — rotor failure at high speed is catastrophic.

Filtration Principles

Filtration separates particles by passing a liquid through a porous medium. The pore size determines what passes through:

• Depth filtration: particles are trapped within a fibrous matrix (e.g., Whatman filter paper). Used for coarse clarification.
• Membrane filtration: a thin membrane with defined pore size (0.2–10 µm) retains particles on the surface. Used for sterilization, particle removal.
• Ultrafiltration: membranes with pores in the nanometer range (1–100 nm) retain macromolecules while allowing small molecules and solvent to pass. Used for protein concentration and buffer exchange (centrifugal concentrators).
• Microfiltration (0.1–10 µm): removes bacteria, yeast, and fine particulates.

Syringe Filters and Filter Units

Syringe filters with 0.2 µm or 0.45 µm membranes are used for small-volume sterilization of samples before HPLC or cell culture. Vacuum-driven filter units (bottle-top filters) handle larger volumes. Always pre-wet membranes for aqueous solutions and confirm chemical compatibility with the solvent.