Question

How does size exclusion column work and what are the rate limiting factors in purification method?

Answer 1

Size exclusion column is a type of chromatographic separation of larger molecules such as proteins

It works on the basis of size or molecular weight of the molecules by the process of filtration

Method:-

• Chromatography column is packed with fine porous beads which are made up of dextran or agarose
• Aqueous solution which which contains molecules to be separated are used to transport the sample column
• Molecules get separated acgieufnati their size
• Small molecules diffuse into the pores and large molecules do not diffuse Into the pores
• And thus separation is achieved

Rate limiting factor in purification method are

• Length of the column:- column length is directly proportional to column capacity. Which means that increase in column length increases the column capacity and the filtration can be achieved efficiently
• Viscosity of the sample:- viscosity measures the flow of the sample.moderate flow is essential for proper separation
• Type of the gel used for separation
• Fineness of the pores in the column for smaller molecules to pass through it.

Hope the answer is clear to you.. thank you!!