10 A Separation and Clarification Flashcards
What is the first step for an extracellular bioproduct?
- Remove / separate liquid material (supernatant)
What is the first step for an intracellular bioproduct?
- Remove / separate solid material (cell mass)
Separation
Means of separating solid and liquid material
ex:
- settling chambers
- centrifugation
- filtration
Settling Chamber
use gravity to remove heavy/large particles (from liquid or air)
Centrifugation
An instrument that generates centrifugal force.
Commonly used to separate particles in a liquid from the liquid
Clarification
- May also be referred to as Isolation
> precipitation
> filtration
Precipitation
Addition of chemical reagent that will result in a macromolecule precipitating out of solution
Filtration - Membrane
Separation of particles from liquid by applying pressure to the solution to force the it through a membrane/filter.
Membrane filtration systems are composed of four primary components:
- Feed/flow
- Pressure
- Membrane
- Filtrate
Normal Flow Filtration (NFF)
- Traps contaminants larger than the pore size on the top surface of the membrane
- Used for critical applications such as sterilizing and final filtration
Depth Filtration
Entraps contaminants both within the matrix and on the surface of the filter media
Tangential Flow Filtration
- Uses a cross flow (i.e. tangential flow) to reduce build up of retained components on the membrane surface
- Preferred for large scale operations
- Used in:
>Clarification/Purification
>Concentration
>Buffer Exchange
Filtration + Centrifugation
Tube with a built in membrane that uses centrifugal force to filter materials
Peristaltic Pump
Creates a gentle squeezing action to move fluid through flexible tubing
Cell Disruption
- Bacteria often express the protein into internal inclusion bodies
- Cell disruption is required when your expression system does not express the recombinant protein directly into the media
