Table of Contents
Chromatography ;
Chromatography is a technique used to separate the components of a mixture. The mixture is dissolved in a solvent and then passed through a column of material that separates the components based on their chemical properties. The components are then eluted from the column and collected in separate containers.
Molecular Exclusion Chromatography/ Exclusion Chromatography
Molecular exclusion chromatography is a type of chromatography that uses a porous membrane to separate molecules based on size. The molecules are dissolved in a solvent and passed through the membrane. Larger molecules are unable to pass through the pores in the membrane and are separated from the smaller molecules. The smaller molecules are then collected and the larger molecules are rejected.
Gel Exclusion Chromatography
Gel exclusion chromatography is a technique used to separate molecules based on their size. The molecules are placed in a gel, and then a solvent is passed through the gel. The smaller molecules move through the gel faster than the larger molecules, and they are separated from the larger molecules.
Principle of Gel Exclusion Chromatography
Gel exclusion chromatography is a technique used to separate molecules based on their sizes. In this technique, a gel is placed in a column and the molecules are placed in the gel. The molecules are then forced to move through the gel by applying pressure. The larger molecules are unable to move through the gel as quickly as the smaller molecules and are therefore separated from the smaller molecules.
Methods of Gel Filtration Chromatography
The methods of gel filtration chromatography include the use of a gel matrix such as agarose, polyacrylamide, or cellulose derivatives such as Sephacel, Superdex, and Sepharose. The gel is used to separate molecules on the basis of size and shape.
The molecules to be separated are placed in a sample tube, which is then placed in a holder on the top of the gel column. The column is then filled with a buffer solution, typically a salt solution such as sodium chloride or potassium phosphate. The solution is circulated through the column by means of a pump, and the molecules are separated by size and shape as they pass through the gel.
The larger molecules take longer to pass through the gel than the smaller molecules, and so they are eluted from the column first. The smaller molecules are eluted from the column last. This allows the separation of the molecules to be controlled by the size and shape of the molecules, rather than by the rate at which they can diffuse through a solution.
The most common use of gel filtration chromatography is to separate proteins on the basis of their size. The technique can also be used to separate other types of molecules, including carbohydrates, lipids, and nucleic acids.
Application of Gel Filtration Chromatography
Gel filtration chromatography is used for the separation of proteins and other large molecules. The proteins are separated by their size, with the smaller proteins passing through the gel more quickly than the larger proteins. This allows for the separation of proteins with similar sizes.
Disadvantages of Gel Permeation chromatography
The disadvantages of gel permeation chromatography include:
1. It is a relatively slow technique.
2. It is not very efficient at separating molecules that are very similar in size.
3. It can be expensive to set up and operate.
