Size exclusion chromatography – Bio-works Technologies AB

Size exclusion chromatography (SEC)

Size Exclusion Chromatography (Gel Filtration) is a simple and reliable technique for separation of molecular components according to their size. The technique relies on a separation media of porous beads, packed closely together in a column. The column is flushed through with a usually aqueous buffer, also known as the mobile phase, and then the sample is loaded.

The larger molecules of the sample are too big to enter the pores so they flow quickly down the column via the spaces around the beads and elute first. Medium-sized molecules are able to enter the pores but their relative size means that the speed in which they move into and out of the pores is fairly rapid. Consequently they elute next. The smaller-sized molecules are able to easily enter the pores and pass through the interior of the beads. This slower progress results in elution nearer the end of the procedure. The larger the molecule the faster it generally will tend to pass through the column.

This technique tends to be the method of choice for the last, or polishing step, in chromatographic purification methodologies, as it is limited by its slow flow rates and the relatively small sample loads that can be applied to the chromatography column.

Separation ranges

ProductSeparation range (kDalton)Exclusion limit (kDalton)
WorkBeads 40/100 SEC10-150150
WorkBeads 40/1000 SEC50-12001200
WorkBeads 40/10 000 SECVery large molecules and particles>10 000


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