CIM® non-cGMP Epoxy 800 mL Tube
About CIM® Epoxy-800 Tube Monolithic Columns
The CIM® Epoxy-800 Tube Monolithic Column consists of a unique polymeric monolith encased in a specially designed housing. These columns are used for direct coupling of proteins, peptides and other ligands containing amino or thiol groups. The design of these columns allows the mobile phase to run in a radial direction from the outer to the inner surface. In this way, resolution is preserved during scale-up while maintaining the short separation layer length. This short separation layer length and the specially engineered highly porous structure allow operations at elevated flow rates with low-pressure drop (only a few bar).The following information is being provided to ensure proper product care and maximal product life. All other information can be found in the Instruction Manual that accompanied the product.
| Catalogue number: | 813.7175 |
| Tube chemistry: | epoxy |
| Ligand density: | 4.22 ± 0.02 mmol/g dry support |
| Support matrix: | poly (glycidyl methacrylate-co-ethylene dimethacrylate) |
| Tube dimensions: | outer diameter: 105 mm; inner diameter: 65 mm; length: 150 mm; dry bed volume: 800mL |
| Working flow rates: | up to 2000 mL/min (uav = 300 cm/h) |
| Working system pressure: | up to 30 bar (3 MPa); WARNING: Do not exceed the maximum allowed pressure as this might seriously damage your column! |
| Temperature stability: | 4 °C (39 °F) to 50 °C (122 °F) WARNING: Avoid prolonged use at elevated temperatures! |
| Recommended pH: | coupling range 1–14 working range 1–14 WARNING: The working range is only valid for the matrix itself. After coupling, it depends on the stability of the coupled ligand! |
Coupling on CIM® Epoxy-800 Tube Monolithic columns
Coupling the ligand
Coupling is performed at a higher pH where the epoxy groups are more reactive. The upper pH value is limited by the ligand stability. Since this is specific for any particular ligand, it should be thoroughly checked and optimized from case to case. In our experience, coupling at pH 8.0 is a good compromise.Temperature dependence
The coupling of the ligand to the CIM® Epoxy-800 Tube Monolithic Column is obtained by a covalent reaction between the epoxy groups on the support and the amino or thiol groups of the ligand. Therefore, the coupling rate increases exponentially with temperature according to the Arrhenius equation. From this point of view, an elevated temperature would be preferred for shortening the coupling time. On the other hand, many ligands may become unstable at higher temperatures and tend to agglomerate or even to loose their biological activity. The coupling temperature depends on the stability of your ligand and should be optimized from case to case. For most ligands (especialy those with high molecular masses) the coupling is best performed for several days at 4 °C. If, however, the ligand is stable, room or even higher temperature is recommended.Buffer composition
According to our experience, the buffer type does not significantly influence the coupling efficiency. Of course, in some cases, the selection of the buffer is of utmost importance, especialy when the ligand stability depends on the environment. Because of this, physiologic buffers are prefferred. Tris, glycine or other nucleophilic components should be avoided as they may compete with the ligand for the epoxy groups, thus lowering the coupling efficiency. For coupling, the ligand should be dissolved in a coupling buffer. In some cases, it is necessary to add organic solvents to dissolve the ligand. The polymeric backbone of the CIM® monolith is hydrophobic. Because of this, coupling can be enhanced by using buffers with higher ionic strengths (e. g. 0.5 M). In this way, the ligand comes closer to the surface facilitating coupling. Higher ionic strengths may not be appropriate as some ligands can start to agglomerate and hinder coupling.Blocking the excess epoxy groups
After the immobilization, the remaining epoxy groups must be blocked in order to avoid unspecific interactions during use. The epoxy groups can be blocked with various reagents like e. g. 1M ethanolamine, overnight at 25–35 °C, 0.1M mercaptoethanol, pH 8, overnight at 25–35 °C, or 0.5 M sulfuric acid, for about 3 hours at 50 °C.WARNING: The selection of a suitable reagent for blocking the excess epoxy groups depends on the coupled ligand's stability!
Storage of CIM® Epoxy-800 Tube Monolithic Columns
CIM® Epoxy-800 Tube Monolithic Column should be stored at a temperature of +4 °C to +8 °C in the presence of 20% ethanol.LOOK-UP CHEMISTRY
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