TSKgel UP-SW3000
TSKgel UP-SW3000 silica-based GFC columns are packed 2 μm particles, providing high efficiency UHPLC separations of proteins. The 25 nm pore size results in a fractionation range up to 500,000 Da for globular proteins. These UHP-SEC columns are ideally suited for the analysis of antibodies, their fragments, and aggregates in biopharmaceutical R&D or QC analysis. They feature the same pore size as TSKgel SWXL, the current industry’s gold standard for QC analysis of therapeutic antibodies. Thus, the UP-SW UHPLC columns allow a smooth method transfer from HPLC to UHPLC. For combination with multi-angle light scattering (MALS) or mass spectrometry (MS) a dedicated column is available: TSKgel UP-SW3000-LS.
It is important to employ an UHPLC system to take full advantage of the high efficiency that can be obtained on TSKgel UP-SW columns. The columns can also be used with an HPLC system that is optimized with regards to extra-column void volume but maximum plate numbers will not be achieved with HPLC instruments.
Protein calibration curve for TSKgel UP-SW3000 and competitor SEC column
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Column: TSKgel UP-SW3000, 2 µm, 4.6 mm ID × 15 cm
Competitor SEC, 1.7 µm, 4.6 mm ID × 15 cm
Mobile phase: 100 mmol/L phosphate buffer, pH 6.7
+ 100 mmol/L Na2SO4 + 0.05% NaN3
Flow rate: 0.35 mL/min
Detection: UV @ 280 nm
Temperature: 25 °C
Inj. vol.: 5 μL
Samples: 1. thyroglobulin, 640 kDa
1’. thyroglobulin dimer
2. gamma-globulin, 155 kDa
2’. gamma-globulin dimer
3. ovalbumin, 47 kDa
4. ribonuclease A, 13,700 Da
5. p-amino benzoic acid, 137 Da
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Comparison of Antibody Analysis Results
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mouse-human chimeric mAb
1: trimer; 2: dimer; 3: monomer ; 4: fragment
| Column |
RS (peak 1/2) |
RS (peak 2/3) |
| TSKgel UP-SW3000 2 μm |
1.52 |
3.56 |
| Competitor UHPLC-SEC 1.7 μm |
1.25 |
3.47 |
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TSKgel UP-SW3000 column-to-column reproducibility
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Column: TSKgel UP-SW3000, 2 µm, 4.6 mm ID × 30 cm
Instrument: Thermo Fisher/Dionex Ultimate 3000 UHPLC System
Mobile phase: 100 mmol/L sodium phosphate buffer, pH 6.7, +
100mmol/L Na2SO4 + 0.05% NaN3
Gradient: Isocratic
Flow rate: 0.35 mL/min
Detection: UV @ 280 nm
Temperature: 25 °C
Injection vol.: 5 µL
Samples: QC standard protein test mixture:
thyroglobulin, 640 kDa, 0.5 g/L
gamma-globulin, 155 kDa, 1 g/L
ovalbumin, 47 kDa, 1 g/L
ribonuclease A , 13,700 Da, 1.5 g/L
p-aminobenzoic acid, 137 Da, 0.01 g/L |