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Education Centre
About the purification of biomolecules
Purpose of purification
Developing purification protocols
How to combine purification steps
Purification development - summary
LC techniques
Affinity Chromatography
Desalting & Gel Filtration
Animation
Basic Principles
The separation mechanism
The gel filtration experiment
Separation modes
Characterisation of gel filtration media
Resolution in Gel filtration
Gel filtration in practice
Technique Profile
What is Gel Filtration?
Hydrophobic interaction chromatography
Ion exchange chromatography
Reversed phase chromatography
Protein Purifier software
BioProcess™ Glossary
The gel filtration experiment

The typical gel filtration experiment consists of three phases as illustrated below:

Explanation of the symbols

Symbolic representation of a
section of a gel filtration bead
surface. The light parts represent
pores of various sizes.

    Sample molecule with no access to the pores.
    Sample molecule with partial access to the pores (the larger ones in the bead section!)
    Sample molecule with full access to the pores.

.
1. Equilibration.
The column is equilibrated with the buffer intended for the separation.


2. Sample application.
The sample is added as a sharp narrow zone at the top of the column. For good resolution the sample volume should be restricted to 0.5 - 5% of the column volume.
3. Elution.
Elution buffer (normally the same type buffer as that used for equilibration) is pumped through the column to make the sample components separate by continuous re-partitioning.
.