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咨询Q NUPharose series is a type of strong anion exchanger resins. Q represents the quaternary ammonium group, NUPharoseTM is the classic agarose gel chromatography matrix base bead of ABSIN. Compared with other Q resins on the market, this product has characteristics such as low non-specific adsorption and a relatively small impact of residence time on the dynamic binding capacity. It is widely used in steps such as capture, intermediate purification, and final purification in the separation and purification of biological macromolecules.
According to differences in resolution and applicable scenarios, Q NUPharose series can be divided into three categories: Q NUPharose FF (~90 μm), Q NUPharose HP (~35 μm), and Q NUPharose BB (~200 μm). FF, which means Fast Flow, endows Q NUPharose FF with the feature of fast flow rate, enabling it to be used for rapid protein purification, have a wide range of applicability and be easy to scale up. HP, which means High Performance, indicates that Q NUPharose HP, with the characteristic of high resolution, is mostly used in the polishing process. BB, which means Big Beads, indicates that Q NUPharose BB, which has a large particle size, is suitable for high-viscosity systems such as blood products and can also be used for batch static operations.
Ion exchange chromatography (IEX) is a separation and purification process in which charged biomolecules exchange ions with IEX resins according to differences in their net surface charge. At a pH above the isoelectric point (PI) of a protein, a protein is negatively charged and can bind to an anion exchanger; while at a pH below the PI of a protein, a protein is positively charged and can bind to cation exchanger. By adjusting the elution conditions (salt concentration and/or pH), different molecules can be eluted in order, thereby achieving the purpose of separation and purification.
After equilibration, the anion exchanger is surrounded by negatively charged counterions, usually Cl-. Oppositely charged proteins bind to anionic groups, becoming concentrated on the column. Uncharged proteins or positively charged proteins pass through the column. For anion exchange chromatography, target molecules can be eluted by increasing the ionic strength of the buffer or lowering the pH (Fig 1). Increasing the ionic strength of the buffer is a competitive elution process. After the ionic strength of the buffer is increased, the negative charge competes with the target molecule to become the counterions of the ion exchanger groups. The proteins with the lowest net charge at the selected pH will be the first ones eluted from the column as ionic strength increases. Similarly, the proteins with the highest charge at a certain pH will be most strongly retained and will be eluted last. After lowering the pH, the net charge of the protein varies.
图1. 阴离子交换填料工作原理示意图
| Product | Q阴离子交换琼脂糖凝胶FF | Q阴离子交换高分辨琼脂糖凝胶HP |
| Matrix | 6% crosslinked agarose | |
| Ligand | quaternary ammonium salt，-CH2CH(OH)CH2N(CH3)3+ | |
| Particle rangea | 45~165 μm | 20~60 μm |
| Particle size, d50 | ~90 μm | ~35 μm |
| Ionic capacity | 110~160 μmol OH-/mL | 60~100 μmol OH-/mL |
| Dynamic binding capacity, DBC10% | ≥40 mg/mLb ≥110 mg/mLc | ≥80 mg/mLb |
| Recommended operating flow velocity | 150~300 cm/h | 60~150 cm/h |
| Maximum flow velocitye | 1200 cm/h< | 300 cm/h |
| Chemical stability | Stable to common aqueous buffers, 1.0 M NaOH, 8 M Urea, 6 M guanidine hydrochloride, 30% isopropanol and 70% ethanol | |
| pH stability | 3~12（Recommended operating, pH），3~12（Long-term）；2~14（CIP） |
注：
a:Microspheres accounting for more than 90% of the volume are in this particle size range.；
b:DBC10% at 10% breakthrough for bovine serum albumin with 2 min residence time；
c:DBC10% at 10% breakthrough for Recombinant Human Epidermal Growth Factor (rh-EGF, ABSIN, NRPA11S) with 2 min residence time；
d:2 mg/mL bovine serum albumin incubate for 2 h at 30 °C；
e:1 cm diameter column and 10 cm bed height using buffers with the same viscosity as water at 20 °C.