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Size Exclusion Chromatography

Size Exclusion Chromatography

Size Exclusion Chromatography (SEC), also referred to as gel-filtration chromatography when used with aqueous mobile phases, uses porous particles to separate biological molecules of different sizes. The advantages of this method include good separation of large molecules from the small molecules while preserving the biological activity of the particles to be separated.

Proteins larger than the pore size cannot enter the pores and will thus co-elute as the first peak in the chromatogram. These molecules are said to be totally excluded from the pores. Proteins that at least partially enter the pores will have varying residence times dependent on their hydrodynamic radius. Different proteins will therefore have different residence times when passing through the column. Molecules that are smaller than the pore size are able to enter all of the pores in the resin and will therefore have the longest residence times. These molecules typically elute together as the last peak in a chromatogram.

 

Applications

The main applications of size exclusion chromatography are desalting of the feedstock, removal of aggregates in a final polishing step and virus purification. In general it can be used to fractionate and purify synthetic and biological polymers, such as proteins, polysaccharides and nucleic acids based on size.  The technique is generally combined with others that further separate molecules by other characteristics, such as pI, charge, and affinity for certain compounds and is typically used as a polishing step. 

 

TOYOPEARL SEC Resins 

TOYOPEARL and TSKgel chromatography media are polymer based macroporous resins for biopurification. Their rigid polymer backbone has better pressure-flow properties the most other commercially available materials. This simplifies column packing and allows for higher operating velocities thereby increasing throughput and decreasing recycling and sanitization times.

TOYOPEARL HW and TSKgel PW size exclusion resins are the foundation for all of Tosoh's ion exchange, hydrophobic interaction, and affinity process media. After an initial polymerization reaction to create the designated particle size and pore size, the resin surface is chemically modified to create the desired final product. Each product is designed for a different molecular weight range and customer feedstock profile.

The table below shows the molar mass separation ranges of each pore size TOYOPEARL HW size exclusion resin:

SEC_MWtable.png


 

TOYOPEARL HW-40 resin is a hydroxylated methacrylic polymer for size exclusion chromatography. It is typically used for high throughput desalting and buffer exchange. It also can remove surfactants such as Triton X100 from protein solutions. It is often used for smaller biomolecules with exclusion limits up to 10,000 Da. 

TOYOPEARL HW-40 resin is available in three particle sizes.

Pore size (mean) 5 nm
Visit the product pages:

TOYOPEARL HW-40EC
TOYOPEARL HW-40C
TOYOPEARL HW-40F
TOYOPEARL HW-40S 

Exclusion limit 10,000 Da
Particle size (mean) 200 µm (EC-grade)
75 µm (C-grade)
45 µm (F-grade)
30 µm (S-grade)
Pressure rating 0.3 MPa
Shipping buffer 20% ethanol
pH stability 2-13
Shelf life 10 years

 

TOYOPEARL HW-50 resin is a hydroxylated methacrylic polymer for size exclusion chromatography of proteins between 500-80,000 Da.

TOYOPEARL HW-50 resin is available in two particle sizes.

Pore size (mean) 12.5 nm Visit the product pages:

TOYOPEARL HW-50F
TOYOPEARL HW-50S
 
Exclusion limit 80,000 Da
Particle size (mean) 45 µm (F-grade)
30 µm (S-grade)
Pressure rating 0.3 MPa
Shipping buffer 20% ethanol
pH stability 2-13
Shelf life 10 years

 

TOYOPEARL HW-55 resin is a hydroxylated methacrylic polymer bead for size exclusion chromatography of proteins between 1,000-700,000 Da. It also is the base bead for all TOYOPEARL "-550" designated products.

TOYOPEARL HW-55 resin is available in two particle sizes.

Pore size (mean) 50 nm Visit the product pages:

TOYOPEARL HW-55F
TOYOPEARL HW-55S
 
Exclusion limit  700,000 Da
Particle size (mean) 45 µm (F-grade)
30 µm (S-grade)
Pressure rating  0.3 MPa
Shipping buffer 20% ethanol
pH stability 2-13
Shelf life 10 years

 

TOYOPEARL HW-65 resin is a hydroxylated methacrylic polymer bead for size exclusion chromatography of proteins between 40,000-5,000,000 Da. It also is the base bead for all TOYOPEARL "-650" designated products.

TOYOPEARL HW-65 SEC resin is available in two particle sizes.

Pore size (mean) 100 nm Visit the product pages:

TOYOPEARL HW-65C
TOYOPEARL HW-65F
TOYOPEARL HW-65S
Exclusion limit 5,000,000 Da
Particle size (mean) 75 µm (C-grade)
45 µm (F-grade)
30 µm (S-grade)
Pressure rating 0.3 MPa
Shipping buffer 20% ethanol
pH stability 2-13
Shelf life 10 years

 

TOYOPEARL HW-75 resin is a hydroxylated methacrylic polymer bead for size exclusion chromatography of proteins between 500,000-50,000,000 Da.

TOYOPEARL HW-75 resin is available in two particle sizes.

Pore size (mean) > 100 nm Visit the product pages:

TOYOPEARL HW-75F
TOYOPEARL HW-75S

Exclusion limit 50,000,000 Da
Particle size (mean) 45 µm (F-grade)
30 µm (S-grade)
Pressure rating 0.3 MPa
Shipping buffer  20% ethanol
pH stability 2-13
Shelf life 10 years