TOSOH SEC
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SEC (Size Exclusion Chromatography) Column
크기 배제 크로마토그래피(SEC)는 겔 여과 크로마토그래피라(GFC)와, 겔 침투 크로마토그래피(GPC)라고도 합니다.
*SEC = GFC (수용성 용매 사용) + GPC (유기용매 사용)
SEC는 다공성 입자를 사용하여 크기가 다른 물질의 분자를 분리합니다.
Pore Size보다 큰 물질은 구멍으로 들어갈 수 없게 되고, 크로마토그램에서 첫 번째 피크로 용출되게 됩니다.
공극 크기(Pore Size)보다 작은 분자는 입자 내 구멍으로 들어갈 수 있기 때문에, 입자로 들어갈 수 없는 큰 분자보다 경로 길이와 체류 시간이 더 깁니다. 이 때, 큰 단백질 분자들은 Pore에서 완전히 배제되었다고 말하고, 부분적으로 공극(Pore)에 들어갈 수 있는 물질은 각각의 개별 물질의 무게와 유체역학적 반지름에 따라 다양한 체류 시간을 가질 것입니다.
따라서, 분자량이 큰 시료는 가장 먼저 용출 되어 크로마토그램에서 첫 번째 피크로 나타나게 되고, 그 뒤로 분자량이 크기 순서대로 용출 되게 됩니다.
Proteins
The TSKgel SW-type columns, first introduced in 1977, are the standard in gel filtration chromatography. GFC is popular among biochemists for the isolation of proteins, for the removal of aggregates, to desalt a protein sample, to separate nucleic acid fractions, or to characterize water-soluble polymers used in food products, paints, pharmaceutical preparations.
The new TSKgel SW mAb products consist of two specialized columns designed for the separation and analysis of monoclonal antibodies (mAb). Compared to competitive columns, these stainless steel, silica-based TSKgel columns offer reduced lot-to-lot variation, long column life, reduction of unspecified adsorption, and superior recovery of aggregates.
TSKgel SW, SWXL and SuperSW columns are silica-based GFC columns optimized for the separation of proteins and nucleic acids using an aqueous buffer as mobile phase. A hydrophilic diol-type bonded phase shields the silica surface from interacting with protein samples. Three different pore sizes are available.
12.5 nm pore size: for analysis of small proteins and peptides
25 nm pore size: for most protein samples
45 nm pore size: for very large proteins and nucleic acids
Compared to competitive HPLC columns, SW-type columns contain more pore volume per unit column volume, which results in higher MW selectivity or better resolution. The TSKgel BioAssist DS column is designed for the desalting and buffer exchange of proteins and polynucleotides at semi-preparative scale.
Particle sizes for the four SW-type and the BioAssist DS column lines are:
TSKgel SuperSW: 4 micron
TSKgel SWXL : 5-8 micron
TSKgel SW: 10-13 micron
TSKgel SW mAb: 3-4 micron
TSKgel BioAssist DS: 15 micron
See each of the TSKgel SW-type and the TSKgel BioAssist DS product pages above for more information and representative calibration curves.
Water-soluble polymers
TSKgel PW columns are the columns of choice for analyses of water-soluble polymers. These TSKgel columns are polymer-based gel filtration columns and prepared from polymethacrylate resin.
TSKgel PW columns exhibit a much larger separation range, better linearity of calibration curves, and less adsorption than the TSKgel Gel Filtration Chromatography SW columns, making them an excellent choice for water-soluble polymers.
While a TSKgel SW column is typically the first column to try for biopolymers, TSKgel PW columns have demonstrated good results for smaller peptides (<1,000 Da), protein aggregates, DNA fragments, and viruses.
TSKgel PW columns: for when analysis time is not critical, when sample mass is not limited, to collect fractions, or to obtain maximum number of plates (at the expense of analysis time).
TSKgel PWXL columns: high efficiency for optimal resolution, reduced analysis time, and in sample limited applications.
TSKgel PWXL-CP columns: chemically modified to improve recovery of cationic polymers.
TSKgel SuperPW columns: semi-micro columns within series allow wide separation range from high MW polymers to oligomers. SuperMultiporePW series are packed with particles containing a wide range of pore sizes.
See each of the TSKgel PW-type product pages for more information and representative calibration curves.
Polar Organic-soluble polymers
TSKgel Alpha and TSKgel SuperAW columns were developed for the GPC analysis of polymers of intermediate polarity. As in the TSKgel PW- and PWXL-type resins, the particles in TSKgel Alpha and SuperAW columns have a hydroxylated methacrylate polymer backbone, but they are crosslinked to a higher degree than TSKgel PW- and PWXL-type columns to minimize swelling in polar organic solvents.
Applications include polymers that are soluble in methanol, acetonitrile, DMSO, isopropanol, or THF. These columns can also be used for water-soluble polymers.
TSKgel SuperAW columns: 4-9 μm particle size
TSKgel Alpha columns: 7-13 μm particle size
TSKgel Super AW and TSKgel Alpha columns feature no or minimal swelling in a wide range of polar organic solvents, including methanol, acetonitrile, DMSO, isopropanol, THF, and HFIP. Accurate MW determination and normal retention of polystrene polymers in DMF solvent can be achieved using either column. System peaks from salts in the eluent elute away from the oligomer of interest.
For high thoughput applications, TSKgel SuperAW columns provide equivalent resolution in half the time compared to TSKgel Alpha columns.
See each TSKgel SuperAW and TSKgel Alpha column product pages for further information and representative calibration curves.
Organic-soluble polymers
TSKgel H series GPC columns are recommended for the analysis of organic-soluble polymers and are packed with spherical particles composed of polystyrene crosslinked with divinylbenzene (PS-DVB). This series includes TSKgel HXL, HHR, SuperH, Super HZ, and SuperMultiporeHZ columns. Each line of columns within this series differs in degree of inertness and operating temperature range.
TSKgel SuperH and TSKgel SuperHZ columns were developed for high-throughput GPC applications. Both product lines feature particle sizes down to 3 μm, a column length of 15 cm and column diameters of 6 mm ID and 4.6 mm ID.
TSKgel HXL and TSKgel HHR columns are conventional GPC columns of 7.8 mm ID x 30 cm, while particles sizes are 5 µm and larger for the larger pore size columns.
Tosoh's innovative multi-pore technology, used in TSKgel SuperMultiporeHZ, produces particles that contain a broad range of pore sizes. Columns with an extended linear calibration curve can now be prepared without mixing particles of different pore sizes.
Except for TSKgel SuperMultiporeHZ columns, all other TSKgel H-type column lines contain at least five different pore sizes. Each product line contains columns for the analysis of oligomers to polymers containing molecular weights as high as an estimated 4 x 108Da. All product lines contain a number of linear columns that are prepared by blending particles with different pore sizes.
See each of the TSKgel H-type product pages for more information and representative calibration curves.